Pas de résumé / Chronic mucocutaneous candidiasis (CMC) is seen in human patients with a variety of conditions and refers to recurrent or persistent infection of the skin, nails and/or mucosae by commensal Candida species. Its pathogenesis had long remained elusive, until human genetic studies of rare patients with inherited forms of idiopathic CMC (whether isolated or syndromic), incriminated impaired interleukin (IL)-17A/F immunity. The first genetic etiologies of idiopathic isolated CMC, autosomal dominant (AD) IL-17F and autosomal recessive (AR) IL-17 receptor A (IL-17RA) deficiencies, were reported in 2011 in a multiplex and in a sporadic case, respectively. Using Whole Exome Sequencing (WES), we identified 26 novel patients bearing 15 different homozygous mutations in the IL17RA gene. The mutations identified are either nonsense; missense; frameshift deletions; frameshift insertions; or non-coding essential splice site mutations. Interestingly, 2 alleles encode for surface expressed receptors, whereas all the other tested alleles are not detected at the surface of the patient’s cells (fibroblasts or leucocytes). IL-17RA deficiency is a fully penetrant AR disease, with early onset symptoms, usually within the first year of life. CMC is always present. In addition, 17 patients present with staphylococcal skin infections, and some patients with pyogenic infections of the respiratory tract, including pneumonia. Interestingly, tuberculosis occurred in two unrelated BCG-vaccinated patients. The response to IL-17A and IL-17F homo- and heterodimers is abrogated in fibroblasts, as well as the response to IL-17E/IL-25 in T cells. Human IL-17RA is thus essential for mucocutaneous immunity against Candida and Staphylococcus, but otherwise largely redundant. AR IL-17RA deficiency should be considered in children or adults with CMC, cutaneous staphylococcal disease, or both. In a separate project, I investigated a female child patient born to consanguineous parents who suffered from CMC, recurrent viral infections, disseminated BCG disease and biliary cryptosporidiosis, suggestive of combined immunodeficiency, and who is homozygous for a mutation in REL, encoding the NF-kB protein c-REL. Sanger sequencing confirmed that the patient is homozygous and that both parents are heterozygous for the mutation, consistent with an AR inheritance. The candidate mutation is a nucleotide substitution localized in an acceptor splice site; is not reported in available public databases; and is predicted to be damaging in silico. The mutation disrupts mRNA splicing and is loss-of expression. The patient shows normal counts of lymphoid subsets, with the exception of diminished frequencies of memory CD4+ T, Th2, Th1*, and memory B cells. The patient’s T cells fail to proliferate in response to recall antigens. Naïve CD4+ T cells produce little IL-2 and respond poorly to polyclonal stimulation, a phenotype reverted by exogenous IL-2. Memory CD4+ T cells also produce little amounts of IL-2, and strongly diminished amounts of key effector cytokines (IFN-γ, IL-4, IL-17A and IL-21). The patient exhibited with no detectable specific antibody response following vaccination. Survival and therefore proliferation of naïve B cells are compromised leading to poor generation of plasma cell, and immunoglobulins secretion. The patient shows normal counts of myeloid cells, and frequencies of dendritic cell subsets. IL-12 production is abolished in whole blood in response to BCG+IFN-γ and B-EBV cells in response to mitogens. Although further investigation is needed to fully characterize the patient’s phenotype, these results strongly suggest that the patient suffers from AR complete c-REL deficiency.

A sepse é a maior causa de morbidade e mortalidade em pacientes queimados, uma vez que profundas alterações ocorrem na farmacocinética de agentes anti-infecciosos prescritos para o controle das infecções. Além disso, pacientes queimados podem apresentar quadro de infecção por germes da comunidade, numa fase precoce de internação na UTI, devendo receber antimicrobianos que diferem daqueles indicados na infecção sistêmica causada por germes hospitalares. Adicionalmente, na vigência de infecção fúngica, o quadro se torna ainda mais grave para os pacientes queimados de prolongada internação e imunocomprometimento. No presente trabalho deve como objetivo realizar o monitoramento plasmático de fluconazol prescrito aos pacientes com infecção sistêmica fúngica internados na UTI e investigar a farmacocinética para o ajuste do regime de dose no controle da infecção fúngica nos pacientes queimados. Investigaram-se 12 pacientes queimados internados na UTI/ Unidade de Queimados - Divisão de Cirurgia Plástica do HC FMUSP, portadores de infecção fúngica recebendo fluconazol através de infusão. Os pacientes receberam o antifúngico geralmente em associação a outros antimicrobianos para o controle das infecções seguindo a recomendação da CCIH do hospital relativa ao regime de dose empírica inicial do controle de infecção na UTI de Queimados. Realizou-se o monitoramento plasmático do fluconazol através da coleta de amostras sanguíneas de pico e vale. Complementarmente, a critério Clínico, foram colhidas amostras seriadas de sangue (pico, 1ª, 2ª, 4ª, 6ª e vale), totalizando seis coletas, para investigação da farmacocinética do agente que requereu ajuste de dose e individualização de terapia no paciente queimado. As coletas de sangue foram realizadas através de cateter venoso (2 mL/coleta em tubos contendo EDTA sódico) pelo médico intensivista de plantão na UTI; o plasma foi obtido pela centrifugação para análise do fármaco de interesse ou então armazenado no congelador (-80° C) até o ensaio. Previamente à realização da Etapa Clínica, foi realizado no Laboratório a validação do método bioanalítico para quantificação do fluconazol no plasma com base na legislação nacional e boas práticas de laboratório, empregando a cromatografia líquida de alta eficiência (CLAE) com detector ultravioleta. A estatística foi realizada pelo tratamento estatístico com utilização do software GraphPad Instat 4.0., GraphPad Prism 4.0, pela utilização de testes não paramétricos. A modelagem farmacocinética foi realizada através da aplicação do software NonCompartmental Analysis, PK Solutions 2.0, aos pares de dados (C vs T) para o agente antifúngico investigado. Os pacientes queimados incluídos no protocolo eram adultos de ambos os sexos 8M/4F, 46,8 ± 20,6 anos, 69,9 ± 11,5 kg, 38,8 ± 24,0% de superfície corporal queimada, e os agentes da queimadura foram para 10 pacientes/ térmico-fogo e para dois pacientes/trauma elétrico; a lesão inalatória foi registrada em 50% pacientes com queimadura pelo fogo. Foram realizados 31 seguimentos farmacoterapêuticos com a emissão de laudos de resultado de exame para o fluconazol. Registrou-se alta variabilidade na farmacocinética para todos os parâmetros investigados. Adicionalmente, registrou-se alteração significativa na farmacocinética do fluconazol nos diversos seguimentos realizados nos pacientes queimados com disfunção renal dialítica quando comparados aqueles em que se registrou função renal preservada. O método bioanalítico mostrou-se adequado ao monitoramento das concentrações plasmáticas do fluconazol através cromatografia líquida de alta eficiência com detecção UV. Registrou-se alta variabilidade na farmacocinética desse agente que justificou em parte a substituição da terapia empírica inicial pela dose ajustada para garantia de terapia eficaz ao paciente queimado. / The sepsis is a main cause of morbidity and mortality in burn patients, once pharmacokinetics of anti-infective drugs prescribed for the control of systemic infections are significantly altered in those patients. In addition, burn patients in the ICU, initially can present infections by community microbial and must receive different antimicrobials than those prescribed for sepsis. On the other hand, burn immunocompromized patients with prolonged staying in the ICU, re-incidence of sepsis and fungal infection requires an effective antifungal agent that must be associated to the antimicrobials prescription. The objective of the study was to therapeutic plasma monitoring of fluconazole largely prescribed to burn patients from the ICU with fungal infection, Pharmacokinetic modeling for dose adjustment and for the control of infection. Twelve burn inpatients with systemic fungal infections from the ICU Burns- Division of Plastic Surgery of Clinics Hospital Medical School University of Sao Paulo received systemically antimicrobials/ antifungal agents. In general burn patients received several antimicrobial agents as recommended by the Control of Hospital Infection Committee as empirical dose at the beginning of therapy and also afterwards in the ICU. The control of infections by community microbial or yet by hospital microbial, and also for fungal infection, was performed by drug plasma after blood sample collection at the peak and at the trough. Complementary, usually by clinical criteria, six blood sample collections were performed at time dose interval (end of drug infusion 1st, 2nd, 4th, 6th and at the trough) for pharmacokinetic purposes, dose adjustment and individualization of drug therapy for burn patients. Blood sample collection was done by the physician from the ICU by venous catheter (2mL/each into blood collection tubes sodium EDTA); plasma obtained by centrifugation of blood tubes were analyzed in the same day or in a deep freezer to storage (-80° C) until assay. Bioanalytical method reported previously was re-validated as recommended by good laboratory practices (GLP). Parameters as linearity, sensitivity, precision and accuracy, recovery and stability, specificity and selectivity were determined for all drugs investigated to guaranteed drug plasma measurements during the pharmacotherapeutic follow up in the ICU. Descriptive statistics was performed by applying the software GraphPad Instat 4.0., GraphPad Prism 4.0 non parametric tests. Pharmacokinetics was estimated by applying the software NonCompartmental Analysis, PK Solutions 2.0, to data (C vs T) for each antimicrobial agent. Burn patients included in the protocol were of both genders 8M/4F, 46.8+/- 20.6 yrs, 69.9+/- 11.5 kg, 38.8+/-24.0% TBSA; agents of the accident were fire/ alcohol for 10 patients and electrical trauma for two patients; inhalation injury were described for 50% of patients with fire. High pharmacokinetic variability was registered for the antifungal agent investigated. In addition, significant changes on pharmacokinetic parameters were described for fluconazole for burn patients with dialytic renal dysfunction compared to those with renal function preserved. Bioanalytical methods validated with basis on good laboratory practices (GLP), recommended by the national and international guidelines were adequate for drug plasma monitoring by liquid chromatography, UV detection. High pharmacokinetic variability was obtained for fluconazol. In conclusion, it was demonstrated that PK modeling of antimicrobial is an important tool for the control of severe systemic fungal infection in burn patients.

A sepse é a maior causa de morbidade e mortalidade em pacientes queimados, uma vez que profundas alterações ocorrem na farmacocinética de agentes anti-infecciosos prescritos para o controle das infecções. Além disso, pacientes queimados podem apresentar quadro de infecção por germes da comunidade, numa fase precoce de internação na UTI, devendo receber antimicrobianos que diferem daqueles indicados na infecção sistêmica causada por germes hospitalares. Adicionalmente, na vigência de infecção fúngica, o quadro se torna ainda mais grave para os pacientes queimados de prolongada internação e imunocomprometimento. No presente trabalho deve como objetivo realizar o monitoramento plasmático de fluconazol prescrito aos pacientes com infecção sistêmica fúngica internados na UTI e investigar a farmacocinética para o ajuste do regime de dose no controle da infecção fúngica nos pacientes queimados. Investigaram-se 12 pacientes queimados internados na UTI/ Unidade de Queimados - Divisão de Cirurgia Plástica do HC FMUSP, portadores de infecção fúngica recebendo fluconazol através de infusão. Os pacientes receberam o antifúngico geralmente em associação a outros antimicrobianos para o controle das infecções seguindo a recomendação da CCIH do hospital relativa ao regime de dose empírica inicial do controle de infecção na UTI de Queimados. Realizou-se o monitoramento plasmático do fluconazol através da coleta de amostras sanguíneas de pico e vale. Complementarmente, a critério Clínico, foram colhidas amostras seriadas de sangue (pico, 1ª, 2ª, 4ª, 6ª e vale), totalizando seis coletas, para investigação da farmacocinética do agente que requereu ajuste de dose e individualização de terapia no paciente queimado. As coletas de sangue foram realizadas através de cateter venoso (2 mL/coleta em tubos contendo EDTA sódico) pelo médico intensivista de plantão na UTI; o plasma foi obtido pela centrifugação para análise do fármaco de interesse ou então armazenado no congelador (-80° C) até o ensaio. Previamente à realização da Etapa Clínica, foi realizado no Laboratório a validação do método bioanalítico para quantificação do fluconazol no plasma com base na legislação nacional e boas práticas de laboratório, empregando a cromatografia líquida de alta eficiência (CLAE) com detector ultravioleta. A estatística foi realizada pelo tratamento estatístico com utilização do software GraphPad Instat 4.0., GraphPad Prism 4.0, pela utilização de testes não paramétricos. A modelagem farmacocinética foi realizada através da aplicação do software NonCompartmental Analysis, PK Solutions 2.0, aos pares de dados (C vs T) para o agente antifúngico investigado. Os pacientes queimados incluídos no protocolo eram adultos de ambos os sexos 8M/4F, 46,8 ± 20,6 anos, 69,9 ± 11,5 kg, 38,8 ± 24,0% de superfície corporal queimada, e os agentes da queimadura foram para 10 pacientes/ térmico-fogo e para dois pacientes/trauma elétrico; a lesão inalatória foi registrada em 50% pacientes com queimadura pelo fogo. Foram realizados 31 seguimentos farmacoterapêuticos com a emissão de laudos de resultado de exame para o fluconazol. Registrou-se alta variabilidade na farmacocinética para todos os parâmetros investigados. Adicionalmente, registrou-se alteração significativa na farmacocinética do fluconazol nos diversos seguimentos realizados nos pacientes queimados com disfunção renal dialítica quando comparados aqueles em que se registrou função renal preservada. O método bioanalítico mostrou-se adequado ao monitoramento das concentrações plasmáticas do fluconazol através cromatografia líquida de alta eficiência com detecção UV. Registrou-se alta variabilidade na farmacocinética desse agente que justificou em parte a substituição da terapia empírica inicial pela dose ajustada para garantia de terapia eficaz ao paciente queimado. / The sepsis is a main cause of morbidity and mortality in burn patients, once pharmacokinetics of anti-infective drugs prescribed for the control of systemic infections are significantly altered in those patients. In addition, burn patients in the ICU, initially can present infections by community microbial and must receive different antimicrobials than those prescribed for sepsis. On the other hand, burn immunocompromized patients with prolonged staying in the ICU, re-incidence of sepsis and fungal infection requires an effective antifungal agent that must be associated to the antimicrobials prescription. The objective of the study was to therapeutic plasma monitoring of fluconazole largely prescribed to burn patients from the ICU with fungal infection, Pharmacokinetic modeling for dose adjustment and for the control of infection. Twelve burn inpatients with systemic fungal infections from the ICU Burns- Division of Plastic Surgery of Clinics Hospital Medical School University of Sao Paulo received systemically antimicrobials/ antifungal agents. In general burn patients received several antimicrobial agents as recommended by the Control of Hospital Infection Committee as empirical dose at the beginning of therapy and also afterwards in the ICU. The control of infections by community microbial or yet by hospital microbial, and also for fungal infection, was performed by drug plasma after blood sample collection at the peak and at the trough. Complementary, usually by clinical criteria, six blood sample collections were performed at time dose interval (end of drug infusion 1st, 2nd, 4th, 6th and at the trough) for pharmacokinetic purposes, dose adjustment and individualization of drug therapy for burn patients. Blood sample collection was done by the physician from the ICU by venous catheter (2mL/each into blood collection tubes sodium EDTA); plasma obtained by centrifugation of blood tubes were analyzed in the same day or in a deep freezer to storage (-80° C) until assay. Bioanalytical method reported previously was re-validated as recommended by good laboratory practices (GLP). Parameters as linearity, sensitivity, precision and accuracy, recovery and stability, specificity and selectivity were determined for all drugs investigated to guaranteed drug plasma measurements during the pharmacotherapeutic follow up in the ICU. Descriptive statistics was performed by applying the software GraphPad Instat 4.0., GraphPad Prism 4.0 non parametric tests. Pharmacokinetics was estimated by applying the software NonCompartmental Analysis, PK Solutions 2.0, to data (C vs T) for each antimicrobial agent. Burn patients included in the protocol were of both genders 8M/4F, 46.8+/- 20.6 yrs, 69.9+/- 11.5 kg, 38.8+/-24.0% TBSA; agents of the accident were fire/ alcohol for 10 patients and electrical trauma for two patients; inhalation injury were described for 50% of patients with fire. High pharmacokinetic variability was registered for the antifungal agent investigated. In addition, significant changes on pharmacokinetic parameters were described for fluconazole for burn patients with dialytic renal dysfunction compared to those with renal function preserved. Bioanalytical methods validated with basis on good laboratory practices (GLP), recommended by the national and international guidelines were adequate for drug plasma monitoring by liquid chromatography, UV detection. High pharmacokinetic variability was obtained for fluconazol. In conclusion, it was demonstrated that PK modeling of antimicrobial is an important tool for the control of severe systemic fungal infection in burn patients.

Persteril (Acidum peraceticum) is a trademark used for a disinfectant with peracetic acid as an active ingredient. It is highly effective biocide and has extensive application possibilities with regard to environmental friendliness, it also has the widest range of disinfection efficacy. The practical part took place in the Genetic centre hatchery at the FROV JU in Vodnany. Persteril? was used for Short-time bath followed by rearing roe in the recirculating system and for short-time bath followed by rearing roe in the flow system. Both versions are breeding quite well; Persteril? detects fungal infection of fish roe in comparison with other liquids, as well as it leaves smaller or no residue in the water or out of the water until the evaporation (according to the concentration).

Background: Necrotizing pancreatitis is a life-threatening presentation of acute pancreatitis. The mortality of 20–80% initially depends on the persistence of organ failure and systemic inflammatory response syndrome (SIRS) and, in the later course of the disease, on secondary infection of the necrosis. The questions whether prophylactic antibiotics aiming to prevent this infection should be administered and which antibiotic is the best to use, as well as the problem of fungal infection under antibiotic treatment are still intriguing and insufficiently solved. Methods: A search of the literature using PubMed was carried out, supplemented by a review of the programmes of the Digestive Disease Week (DDW) and the United European Gastroenterology Week (UEGW). Results: Despite the widely practised prophylactic antibiotic administration in severe pancreatitis, no evidence for the benefit of this strategy exists. One of the drawbacks might be a tendency for disastrous fungal infection under prophylactic antibiotics. Bacterial translocation from the gut in the second week after the onset of symptoms is the major source for infection of pancreatic necrosis and provides a clear indication for antibiotic treatment. However, routine fine-needle aspiration for a calculated antibiotic therapy cannot be recommended, and all other tests offer only indirect signs. Important factors such as enteral versus parenteral feeding and the method of necrosectomy are mostly neglected in the trials but seem to be essential for the outcome of the patient. Conclusions: Even though most meta-analyses including the newer double-blind, placebo-controlled trials on prophylactic antibiotics showed no beneficial effects in the prevention of infection of necrosis and/or outcome of the patients, this strategy is still widely used in clinical routine. Since nearly all trials published so far show systematic problems (i.e. inaccurate definition of the severity of the disease, poor statistical testing, and neglect of differences in the route of nutrition), there is a need for randomized controlled prospective trials with exact definitions of the disease. / Hintergrund: Die nekrotisierende Pankreatitis weist eine Mortalität von 20–80% auf. Initial ist vor allem das Ausmaß des Organversagens entscheidend für die Prognose des Patienten. In der zweiten Krankheitswoche stellt dann die sekundäre Infektion der Nekrosen durch die Translokation von Darmkeimen das entscheidende Problem dar. Zur Vermeidung einer solchen Infektion werden klinisch sehr häufig Breitspektrumantibioktika prophylaktisch eingesetzt. Dies wird aber zunehmend kritisch diskutiert, und es existieren kontroverse Empfehlungen. Methoden: Eine Literaturrecherche unter Einbeziehung von PubMed und der Programme der Digestive Disease Week (DDW) und der United European Gastroenterology Week (UEGW) wurde durchgeführt. Ergebnisse: Die meisten Studien können den prophylaktischen Einsatz von Antibiotika bei der schweren Pankreatitis nicht rechtfertigen. Einige Studien belegen vielmehr eine Selektion resistenter Keime und vor allem auch eine erhöhte Rate von schwer therapierbaren Pilzinfektionen unter einer solchen Therapie. Daher sollte erst nach dem Nachweis einer Nekroseinfektion mit einer Antibiotikatherapie begonnen werden, wobei keine Routine-Feinnadelpunktion der Nekrose zum Keimnachweis durchgeführt werden sollte. Es stehen daher nur indirekte, meist bildgebende Verfahren für den Infektionsnachweis zur Verfügung. Entscheidende Faktoren wie die enterale Ernährung und die Methode der Nekrosektomie wurden bisher bei den meisten Studien vernachlässigt, scheinen aber essenziell für das Behandlungsergebnis des Patienten zu sein. Schlussfolgerungen: Die meisten publizierten Studien weisen eine sehr heterogene Definition der Erkrankung, uneinheitliche Behandlungsprotokolle und Ungenauigkeiten bei der statistischen Testung auf. Gerade entscheidende Faktoren wie die enterale Ernährung werden größtenteils komplett vernachlässigt. Es besteht daher ein Bedarf für randomisierte placebokontrollierte Studien, die diese Probleme berücksichtigen und suffiziente Schlussfolgerungen zur Antibiotikatherapie der schweren Pankreatitis zulassen.

The interleukin (IL)-12 family of cytokines plays a key role in the orchestration of cellular immune responses, bridging innate and adaptive immunity. The founding member, IL-12, was discovered in the late 1980s as the first heterodimeric cytokine, composed of a 40 kDa (p40) and 35 kDa (p35) subunit. Years of basic and clinical research on this prototypical T helper type (Th)1 cytokine revealed its importance in immunity to intracellular non-viral infections, as well as in cancer and autoimmune diseases. Since the discovery of IL-23 as another cytokine composed of the p40 subunit of IL-12 in the year 2000, IL-23, rather than IL-12, could be shown to be the key player in rodent models of autoimmune diseases such as multiple sclerosis and rheumatoid arthritis. With accumulating evidence revealing IL-23 as the crucial regulator of a non-classical pathway of cellular immunity which is hallmarked by IL-17 producing T cells it is intriguing to gain understanding of the importance of such findings in immunity to infections. The present work describes a series of in vivo studies investigating the role of endogenous as well as exogenous IL-23 in a murine model of chronic fungal infection, cryptococcosis. To address the role of endogenous IL-23, wild-type (WT), IL-12- (IL-12p35-/-), IL-23- (IL-23p19-/-) deficient, as well as IL-12- and IL-23- double deficient (p40-deficient) mice on a C57BL/6 background were infected with Cryptococcus neoformans (C. neoformans). Following infection, p40-deficient mice demonstrated higher mortality than IL-12p35-/- mice. Reconstitution of p40-deficient mice with recombinant murine IL-23 prolonged their survival to levels similar to IL-12p35-/- mice. IL-23p19-/- mice showed a moderately reduced survival time and delayed fungal clearance in the liver. While interferon (IFN)-γ production was similar in WT and IL-23p19-/- mice, production of IL-17 was strongly impaired in the latter. IL-23p19-/- mice produced fewer hepatic granulomata relative to organ burden and showed defective recruitment of mononuclear cells to the brain. Moreover, activation of microglia cells and expression of IL-1β, IL-6, and MCP-1 in the brain was impaired. SUMMARY - 80 - The second part of the present work explores the mechanisms underlying the IL-23 effects by characterizing the role of exogenous IL-23. C. neoformans-infected C57BL/6 WT mice treated with recombinant murine IL-23 showed significantly prolonged survival time as compared to mock-treated control mice. However, complete survival throughout the observation period (100 days) was only achieved following IL-12 treatment. At day 21 post infection (p.i.) the IL-23-treated mice as well as the IL-12 group had a significantly lower fungal burden in the brain than the control mice. However, while IL-12 treatment was associated with elevated serum levels of the proinflammatory mediators IFN-γ, tumor necrosis factor (TNF)-α and nitric oxide, IL-23-treated animals, although more resistant, developed a Th2 response similar to the control group as measured by serum IgE levels. Further experiments to assess the mechanism of action were based on the finding of reduced fungal burden at the site of infection, the peritoneal cavity, at day 8 p.i. following IL-23 treatment. This microbicidal effect was also seen in p40-deficient as well as in T and B cell deficient (RAG-deficient) mice. Administration of IL-23 led to enhanced recruitment of inflammatory cells, not only of T cells but also cells of the innate immune system such as DCs, natural killer cells and granulocytes to the infected site. Although numbers of macrophages were not altered following IL-23 treatment, co-stimulatory molecules were markedly up-regulated on such cells. The chemokine/cytokine pattern induced by IL-23 treatment was hallmarked by proinflammatory mediators such as MCP-1, IL-1β, IL-6, TNF-α and IL-17, but also the Th2 associated cytokine IL-5. From these results it can be concluded that a non-classical immune pathway induced by IL-23 complements the more dominant role of IL-12 in protection against C. neoformans. This novel immune response is characterized by an enhancement of the inflammatory cell response and the production of a proinflammatory cytokine pattern hallmarked by IL-1β, IL-6, TNF-α and IL-17.

Cryptococcus neoformans, an opportunistic fungal pathogen ubiquitously present in the environment, causes cryptococcal meningitis (CM) mainly in immunocompromised patients, such as AIDS patients. We aimed to identify disease-associated cryptococcal protein antigens targeted by the human humoral immune response. Therefore, we used sera from Colombian CM patients, with or without HIV infection, and from healthy individuals living in the same region. Serological analysis revealed increased titers of anti-cryptococcal IgG in HIV-negative CM patients, but not HIV-positive CM patients, compared to healthy controls. In contrast, titers of anti-cryptococcal IgM were not affected by CM. Furthermore, we detected pre-existing IgG and IgM antibodies even in sera from healthy individuals. The observed induction of anti-cryptococcal IgG but not IgM during CM was supported by analysis of sera from C. neoformans-infected mice. Stronger increase in IgG was found in wild type mice with high lung fungal burden compared to IL-4Ra-deficient mice showing low lung fungal burden. To identify the proteins targeted by human anti-cryptococcal IgG antibodies, we applied a quantitative 2D immunoproteome approach identifying cryptococcal protein spots preferentially recognized by sera from CM patients or healthy individuals followed by mass spectrometry analysis. Twenty-three cryptococcal proteins were recombinantly expressed and confirmed to be immunoreactive with human sera. Fourteen of them were newly described as immunoreactive proteins. Twelve proteins were classified as disease-associated antigens, based on significantly stronger immunoreactivity with sera from CM patients compared to healthy individuals. The proteins identified in our screen significantly expand the pool of cryptococcal proteins with potential for (i) development of novel anticryptococcal agents based on implications in cryptococcal virulence or survival, or (ii) development of an anti-cryptococcal vaccine, as several candidates lack homology to human proteins and are localized extracellularly. Furthermore, this study defines preexisting anti-cryptococcal immunoreactivity in healthy individuals at a molecular level, identifying target antigens recognized by sera from healthy control persons.

Effective prevention, or treatment, of invasive fungal infection in the neutropenic patient has hitherto been unsatisfactory because of either an inadequate anti-fungal spectrum of the agent or important toxicity. Itraconazole is effective against a broad spectrum of the opportunistic pathogens seen in Europe and North America. Prior problems with absorption, e.g. in the marrow transplant recipient, have been overcome with the introduction of an oral solution and an i.v. preparation. The deliberations of an expert meeting held in June, 1998 include recommendations on which patient requires one of these new preparations based on clinical trials, the dose and route. Important drug interactions are also detailed. / Dieser Beitrag ist mit Zustimmung des Rechteinhabers aufgrund einer (DFG-geförderten) Allianz- bzw. Nationallizenz frei zugänglich.

Leveduras da mucosa oral, de cateter e de sangue de neonatos internados na UTIN (Unidade de Terapia Intensiva Neonatal) por período de 9 meses, foram avaliadas quanto ao perfil molecular pela técnica de PFGE. Dos 125 neonatos internados, 23 (18,4%) apresentaram leveduras na mucosa oral, sangue e cateter. Destes pacientes, obtivemos 54 amostras de leveduras, sendo 36 (66,7%) isoladas da colonização oral; 12 (22,2%) de sangue, seis (11,1%) de cateter. C. albicans foi a mais freqüente dentre as amostras isoladas da mucosa oral e sepse. C. parapsilosis foi a mais isolada dentre as amostras de cateter. Todas as amostras foram consideradas sensíveis aos antifúngicos. Estas amostras apresentaram atividade de pelo menos uma exoenzima. Dentre os 12 casos de sepse, 83,4% (10/12) foram causadas por espécies de Candida, sendo que, destas 10 candidemias, 60% (6/10) estiveram associadas com colonização oral prévia pela mesma espécie e perfil genotípico. A mortalidade devido a infecção sistêmica (septicemia) por leveduras,no período do estudo, foi de 91.6% . / Yeasts isolated from the oral mucosa, catheter and blood of neonates in na NICU (neonatal intensive care unit) over a period of 9 months, were were evaluated in regard to the molecular profile by PFGE technique. Of the 125 neonates studied, 23 (18.4%) presented yeast in their oral mucosa, blood, or catheter. From these 23 neonates, 54 samples of yeast were obtained; 36 (63.7%) from oral colonization, 12 (22.2%) from the blood, and 6 (11.1%) from the catheter. The most frequently encountered species among the samples isolated from the oral mucosa and blood was C. albicans; while C. parapsilosis was the species found most frequently among the samples obtained from catheters. All the samples were considered susceptible to antifungal agents tested. The samples presented activity of at least one exoenzyme. Among the 12 cases of sepsis, 10 (83.4%) were caused by species of Candida, and of these 10 cases, 6 (60%) were associated with previous oral colonization by the same species and genotypic profile. The mortality rate due to septicemia by yeasts was 91.6%.

No description available.