Leishmaniasis is protozoan diseases, which is transport into the host during the feeding of sand fly. During the feeding of infected sand flies not only the leishmania but also the sand fly saliva are inoculated into the hosts. Sand fly saliva can strongly affect the response of the immune system. If the host hadn't met sand fly saliva yet, the course of infection is usually worse. In cutaneous leishmaniasis, the lesions developed early, being more destructive and perstiting longer, if not healed. The hosts living in endemic areas of leishmaniasis and the vector hosts are often exposed to feeding uninfected sand flies. To hosts are repeatedly inoculated the sand fly saliva antigen and induced specific cellular and antibody responses. Cellular and antibody responses are different for different hosts, attempts were made most frequently in murine and canine models. In humans, as host sis it difficult to monitor development leishamnia infectipon after previous exposure, that's why in humans mainly it is monitors the levels of antibodies, according to which we can determine the extent of sand fly bited and the risk of transmission of leishmaniasis. The specifity of immune responses against sand fly saliva is important for the testing new type of controlling and healing programs against sand fly and...

<p>Coeliac disease (CD) is a multifactorial disease of the small intestine. In genetically predisposed individuals the, ingestion of cereals leads to a remodulation of the mucosal architecture, and the production of autoantibodies against tissue transglutaminase (tTG). The treatment is a lifelong gluten-free diet.</p><p>The diagnostic procedure relies on the examination of a small-bowel biopsy that displays villous atrophy. A spectrum of clinical manifestations is associated with CD, ranging from overt enteropathy to atypical and silent symptoms. Approximately 1% of the general population has CD, and the majority is undiagnosed. Although most patients with active CD can be detected by the assessment of elevated IgA-tTG, some patients lack these antibodies. Moreover, individuals with IgA-deficiency cannot be identified by means of IgA serology. </p><p>The aim of this thesis was to investigate the clinical utility of IgG-tTG for the detection and follow-up of subjects with active CD. The included studies showed that IgG-tTG was highly prevalent in IgA-deficient and IgA-competent patients with CD, whereas non-CD patients rarely had these antibodies. During a gluten-free diet, IgG-tTG decreased, demonstrating that IgG-tTG can be used to follow the patient’s adherence such a diet. Furthermore, 10% of healthy IgA deficient blood donors had elevated IgG-tTG, indicating that they had silent CD. </p><p>In IgA-competent subjects, high IgG-tTG levels correlated with a severe mode of CD and profound mucosal deterioration, suggesting that IgG-tTG might be involved in the disease progression. Moreover, we found that although a considerable percentage of IgA-competent patients lack IgG-tTG, the presence of these antibodies in conjunction with high levels of IgA-tTG was highly predictive of a severe small-intestine villous atrophy. It was also demonstrated that IgG-tTG normalisation coincided with clinical remission in IgA-competent CD patients on a gluten-free diet. </p>

Coeliac disease (CD) is a multifactorial disease of the small intestine. In genetically predisposed individuals the, ingestion of cereals leads to a remodulation of the mucosal architecture, and the production of autoantibodies against tissue transglutaminase (tTG). The treatment is a lifelong gluten-free diet. The diagnostic procedure relies on the examination of a small-bowel biopsy that displays villous atrophy. A spectrum of clinical manifestations is associated with CD, ranging from overt enteropathy to atypical and silent symptoms. Approximately 1% of the general population has CD, and the majority is undiagnosed. Although most patients with active CD can be detected by the assessment of elevated IgA-tTG, some patients lack these antibodies. Moreover, individuals with IgA-deficiency cannot be identified by means of IgA serology. The aim of this thesis was to investigate the clinical utility of IgG-tTG for the detection and follow-up of subjects with active CD. The included studies showed that IgG-tTG was highly prevalent in IgA-deficient and IgA-competent patients with CD, whereas non-CD patients rarely had these antibodies. During a gluten-free diet, IgG-tTG decreased, demonstrating that IgG-tTG can be used to follow the patient’s adherence such a diet. Furthermore, 10% of healthy IgA deficient blood donors had elevated IgG-tTG, indicating that they had silent CD. In IgA-competent subjects, high IgG-tTG levels correlated with a severe mode of CD and profound mucosal deterioration, suggesting that IgG-tTG might be involved in the disease progression. Moreover, we found that although a considerable percentage of IgA-competent patients lack IgG-tTG, the presence of these antibodies in conjunction with high levels of IgA-tTG was highly predictive of a severe small-intestine villous atrophy. It was also demonstrated that IgG-tTG normalisation coincided with clinical remission in IgA-competent CD patients on a gluten-free diet.

Neste trabalho foi desenvolvido um Dot-ELISA capaz de detectar anticorpos anti-Plasmodium vivax, anti-Plasmodium falciparum, anti-Trypanosoma cruzi e anti-Treponema pallidum, simultaneamente. O teste foi padronizado e aplicado em amostras de pacientes com malária, doença de Chagas e sífilis e comparado com testes de referência. Foi utilizado no estudo 52 amostras de pacientes com doença de Chagas, 43 pacientes com sífilis, 103 indivíduos com infecção presente (primo-infectado e não primo-infectados) ou passada de malária, indivíduos com anticorpos heterólogos, 30 indivíduos com leishmaniose 100 indivíduos saudáveis. O Dot-ELISA-Multi apresentou 100% de especificidade para todos os antígenos nas amostras de indivíduos saudáveis e com anticorpos heterólogos, com exceção do antígeno TESA que obteve 99%. A sensibilidade obtida foi de 100% em indivíduos chagásicos e 88% em pacientes com sífilis. Em indivíduos com malária a sensibilidade obtida foi de 90% para PvMSP119 (antígeno de P. vivax) e 47% para Pf-Zw (antígeno de P. falciparum). A positividade do teste em indivíduos não parasitados com histórico de malária foi de 92%. Nas amostras de malária observou-se que em indivíduos que tinham tido ultimo episódio de P. vivax, associação negativa foi observada entre o tempo passado desde o último episódio e reatividade de Dot-ELISAMulti PvMSP119, e associação positiva entre o número de episódios de malária e reatividade de Dot-ELISA-Multi Pf-Zw. Indivíduos cujo o último episódio foi por P. falciparum, o Dot-ELISA-Multi Pf-Zw apresentou associação positiva com o número de episódios ocorridos. O comparado com os testes de referência utilizados apresentou um nível muito bom de concordância para TESA, EAE, PvMSP119 e um nível bom de concordância para Pf-Zw / In the present study, a Dot-ELISA was assembled to test antibodies against Plasmodium vivax, Plasmodium falciparum, Trypanosoma cruzi and Treponema pallidum and was standardized and evaluated in serum samples from patients with malaria, Chagas disease and syphilis, in comparison with reference tests. The study was carried out on serum samples from 52 patients with chronic Chagas disease, 103 individuals with current (parasitemic) or past malaria (aparasitemic), 43 patients with syphilis, 30 with leishmaniosis, 21 individuals with heterologous antibodies and 100 blood donors. The diagnostic performance of Dot-ELISA-Multi for serum samples from patients with heterologous antibodies and from healthy blood donors, an overall 100% specificity was obtained for all antigens but TESA. A 100% sensitivity was observed in serum specimens from chronic-chagasic patients and 88% in serum specimens from syphilis patients. For malaria samples, the positivity was 90% for PvMSP119 and 47% for Pf-Zw antigen. In past malaria individuals, positivity was 92%. Sera from subjects who had had a P. vivax-malaria last episode presented negative association between time elapsed since their last malaria episode and results from Dot-ELISA-Multi PvMSP119; while positive association was observed between number of malaria episodes and results from Dot-ELISA-Multi Pf-Zw. For individuals who had had a P. falciparum-malaria last episode, Dot-ELISAMulti Pf-Zw results showed positive association with number of malaria episodes only. Altogether, concerning the reactivity of the five antigens of the Dot-ELISAMulti, as compared with their respective reference tests, we have observed a very good level of concordance for TESA, EAE, PvMSP119 and for Tp-Zw and a good level for Pf-Zw

Neste trabalho foi desenvolvido um Dot-ELISA capaz de detectar anticorpos anti-Plasmodium vivax, anti-Plasmodium falciparum, anti-Trypanosoma cruzi e anti-Treponema pallidum, simultaneamente. O teste foi padronizado e aplicado em amostras de pacientes com malária, doença de Chagas e sífilis e comparado com testes de referência. Foi utilizado no estudo 52 amostras de pacientes com doença de Chagas, 43 pacientes com sífilis, 103 indivíduos com infecção presente (primo-infectado e não primo-infectados) ou passada de malária, indivíduos com anticorpos heterólogos, 30 indivíduos com leishmaniose 100 indivíduos saudáveis. O Dot-ELISA-Multi apresentou 100% de especificidade para todos os antígenos nas amostras de indivíduos saudáveis e com anticorpos heterólogos, com exceção do antígeno TESA que obteve 99%. A sensibilidade obtida foi de 100% em indivíduos chagásicos e 88% em pacientes com sífilis. Em indivíduos com malária a sensibilidade obtida foi de 90% para PvMSP119 (antígeno de P. vivax) e 47% para Pf-Zw (antígeno de P. falciparum). A positividade do teste em indivíduos não parasitados com histórico de malária foi de 92%. Nas amostras de malária observou-se que em indivíduos que tinham tido ultimo episódio de P. vivax, associação negativa foi observada entre o tempo passado desde o último episódio e reatividade de Dot-ELISAMulti PvMSP119, e associação positiva entre o número de episódios de malária e reatividade de Dot-ELISA-Multi Pf-Zw. Indivíduos cujo o último episódio foi por P. falciparum, o Dot-ELISA-Multi Pf-Zw apresentou associação positiva com o número de episódios ocorridos. O comparado com os testes de referência utilizados apresentou um nível muito bom de concordância para TESA, EAE, PvMSP119 e um nível bom de concordância para Pf-Zw / In the present study, a Dot-ELISA was assembled to test antibodies against Plasmodium vivax, Plasmodium falciparum, Trypanosoma cruzi and Treponema pallidum and was standardized and evaluated in serum samples from patients with malaria, Chagas disease and syphilis, in comparison with reference tests. The study was carried out on serum samples from 52 patients with chronic Chagas disease, 103 individuals with current (parasitemic) or past malaria (aparasitemic), 43 patients with syphilis, 30 with leishmaniosis, 21 individuals with heterologous antibodies and 100 blood donors. The diagnostic performance of Dot-ELISA-Multi for serum samples from patients with heterologous antibodies and from healthy blood donors, an overall 100% specificity was obtained for all antigens but TESA. A 100% sensitivity was observed in serum specimens from chronic-chagasic patients and 88% in serum specimens from syphilis patients. For malaria samples, the positivity was 90% for PvMSP119 and 47% for Pf-Zw antigen. In past malaria individuals, positivity was 92%. Sera from subjects who had had a P. vivax-malaria last episode presented negative association between time elapsed since their last malaria episode and results from Dot-ELISA-Multi PvMSP119; while positive association was observed between number of malaria episodes and results from Dot-ELISA-Multi Pf-Zw. For individuals who had had a P. falciparum-malaria last episode, Dot-ELISAMulti Pf-Zw results showed positive association with number of malaria episodes only. Altogether, concerning the reactivity of the five antigens of the Dot-ELISAMulti, as compared with their respective reference tests, we have observed a very good level of concordance for TESA, EAE, PvMSP119 and for Tp-Zw and a good level for Pf-Zw

Para elucidar os mecanismos envolvidos na inibição da hipersensibilidade do tipo I em proles murinas mediada pela imunização materna com Ovalbumina (OVA), as proles foram avaliadas quanto aos linfócitos T&#947;&#948; produtores de IL-17 (CD27-). A imunização materna com OVA reduziu a expressão de CD27, o que também se refletiu nos pulmões, e a produção de IL-17 por linfócitos T&#947;&#948; das proles. A redução da expressão de CD27 também foi evidenciada em linfócitos T&#947;&#948; intratímicos das proles após a transferência passiva de anticorpos IgG alérgeno específicos para fêmeas gestantes não imunizadas e in vitro em resposta a estes mesmos anticorpos, efeito que parece envolver a expressão de receptores para IgG expressos por outras células presentes no timo. As evidências obtidas indicam que a imunização materna influi na maturação tímica de linfócitos inibindo a população que colaborara com a inflamação alérgica. Aparentemente, os anticorpos IgG maternos são responsáveis por este fenômeno e a população estudada está envolvida na inibição da inflamação alérgica observada nas proles. / To elucidate the mechanisms involved in inhibition of type I hypersensitivity in murine offsprring mediated by maternal immunization with Ovalbumin (OVA), the offspring were evaluated as IL-17-producing &#947;&#948;T cells (CD27-). Maternal immunization with OVA reduced the expression of CD27, which was also reflected in the lungs, and the production of IL-17 by &#947;&#948;T cells of offspring. Reduction of CD27 expression was also evidenced in intrathymic &#947;&#948;T cells of offspring after passive transfer of allergen-specific IgG antibodies to non-immunized pregnant females and in vitro in response to these same antibodies, effect that seems to involve the expression of IgG receptors expressed by other cells present in the thymus. Evidence obtained indicates that maternal immunization influences the thymic maturation of lymphocytes by inhibiting the population that collaborate with allergic inflammation. Apparently, maternal IgG antibodies are responsible for this phenomenon and the population studied is involved in the inhibition of allergic inflammation observed in offspring.

Para elucidar os mecanismos envolvidos na inibição da hipersensibilidade do tipo I em proles murinas mediada pela imunização materna com Ovalbumina (OVA), as proles foram avaliadas quanto aos linfócitos T&#947;&#948; produtores de IL-17 (CD27-). A imunização materna com OVA reduziu a expressão de CD27, o que também se refletiu nos pulmões, e a produção de IL-17 por linfócitos T&#947;&#948; das proles. A redução da expressão de CD27 também foi evidenciada em linfócitos T&#947;&#948; intratímicos das proles após a transferência passiva de anticorpos IgG alérgeno específicos para fêmeas gestantes não imunizadas e in vitro em resposta a estes mesmos anticorpos, efeito que parece envolver a expressão de receptores para IgG expressos por outras células presentes no timo. As evidências obtidas indicam que a imunização materna influi na maturação tímica de linfócitos inibindo a população que colaborara com a inflamação alérgica. Aparentemente, os anticorpos IgG maternos são responsáveis por este fenômeno e a população estudada está envolvida na inibição da inflamação alérgica observada nas proles. / To elucidate the mechanisms involved in inhibition of type I hypersensitivity in murine offsprring mediated by maternal immunization with Ovalbumin (OVA), the offspring were evaluated as IL-17-producing &#947;&#948;T cells (CD27-). Maternal immunization with OVA reduced the expression of CD27, which was also reflected in the lungs, and the production of IL-17 by &#947;&#948;T cells of offspring. Reduction of CD27 expression was also evidenced in intrathymic &#947;&#948;T cells of offspring after passive transfer of allergen-specific IgG antibodies to non-immunized pregnant females and in vitro in response to these same antibodies, effect that seems to involve the expression of IgG receptors expressed by other cells present in the thymus. Evidence obtained indicates that maternal immunization influences the thymic maturation of lymphocytes by inhibiting the population that collaborate with allergic inflammation. Apparently, maternal IgG antibodies are responsible for this phenomenon and the population studied is involved in the inhibition of allergic inflammation observed in offspring.

The bacteria Streptococcus pyogenes produces a multitude of proteins which interact with and alter the functions of the host immune system. Two such proteins, Endoglycosidase S (EndoS) and Immunoglobulin G-degrading enzyme from S. pyogenes (IdeS) are able to specifically alter the effector functions of immunoglobulin G (IgG). EndoS is a glycoside hydrolase which removes the conserved <i>N</i>-linked glycan from IgG Fc whereas IdeS is a cysteine protease that cleaves the exible protein hinge of IgG. The activity of both proteins results in the reduced ability of IgG to elicit immune responses through Fc receptor binding and complement activation. Amongst other applications, both EndoS and IdeS are actively being explored as new therapeutics for IgG-mediated autoimmune diseases. Given the therapeutic potential of EndoS and IdeS, experiments were designed to investigate the structural and functional characteristics of these enzymes in an effort to understand their specficity for and activity against IgG. Here, bioinformatic and biophysical characterisation of EndoS identified subdomains outside of the catalytic domain which contribute to glycoside hydrolase activity. The substrate specificity of EndoS was also explored and showed that EndoS hydrolyses a broad range of glycans from the IgG scaffold. EndoS was also shown to have activity against alternative glycoprotein substrates, however, this non-specific activity was negligible in the context of whole serum. The effect of EndoS-mediated deglycosylation on the structure of the IgG Fc domain was explored using both X-ray crystallography and small-angle X-ray scattering. Small angle X-ray scattering was also used to characterise both EndoS and IdeS in complex with IgG Fc. Solution-state models of each complex were produced providing preliminary data towards how these enzymes interact with IgG. Overall, the results presented here contribute to our understanding of these enzymes which is of importance as they go forward into clinical applications.

Background: Post COVID-19 Condition (PCC, also known as long COVID and post-acute sequelae of COVID-19) is a major public health concern with severe and pervasive impacts on physical and mental health. PCC is highly heterogeneous and may manifest as different clusters of symptoms of varying intensity and duration. The etiology of PCC remains uncertain, though several underlying pathophysiological mechanisms, such as cellular damage, inflammatory cytokines, and a hypercoagulable state, are thought to contribute to PCC inception and trajectory. Examination of potential serological markers of PCC, accounting for clinical covariates, may yield emergent pathophysiological insights. Objectives: Primary objectives of this thesis are to 1) Identify key clinical and potential serological predictors of PCC; 2) Acquire clinical and serological data in a large-scale prospective observational study; 3) Assess relationships between PCC and serological markers, accounting for clinical covariates; 4) Systematically review evidence to date on primary observational studies comparing serological response between people with and without persistent symptoms post COVID-19 recovery; 5) Discuss persisting gaps in knowledge and data quality, and propose strategies for resolve. Methods: This thesis is framed around three core efforts: 1) The design of survey questions and study materials, recruitment of participants, and data collection in a large-scale prospective cohort study launched in 2020; 2) The assessment of relationships between pre-defined serological predictors and PCC, accounting for clinical covariates; and 3) A robust rapid review of PCC onset and phenotype as functions of serological markers. Expert opinion was sought to define serological predictors. Clinical predictors were defined a priori based on systematic reviews meeting AMSTAR 2 guidelines. Conclusions: To address objectives, we described efforts to collect clinical and serological data from a large-scale prospective cohort study; identify PCC-cases and infected-controls; assess associations between pre-defined serological predictors (IgG titres targeting SARS-CoV-2 spike (S), nucleocapsid (N), and receiver binding domain (RBD) antigens, and efficient neutralization) and PCC; and synthesized findings from an extensive rapid review on PCC as a function of serological markers. Our multivariate analysis using Stop the Spread Ottawa data is, to our knowledge, the first Canadian study to report the direction and magnitude of association between selected serological predictors (anti-IgG response to S, N, and RBD SARS-CoV-2 antigens, and neutralizing efficiency) and PCC status and impact on quality of life. Finally, we described five potential strategies which may improve the accessibility, quality, and amalgamation of data pertaining to PCC: 1) Fostering comparability between studies to enable synthesis of multiple datasets; 2) Advancing the characterization and consensus on PCC phenotypes; 3) Employing innovative modelling strategies that could potentially yield novel insights; 4) Promoting robust collaboration and knowledge sharing among research teams; and 5) Engaging people with lived experience at all stages of research.

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