Mestrado em Biologia da Relação Parasito-Hospedeiro (IPTSP)

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Avaliação do comportamento metabólico de cisticercos de Taenia crassiceps cultivados in vitro e expostos a condições estressantes
(Universidade Federal de Goiás, 2013-09-06) Andrade, Lilian Cristina Morais de; Vinaud, Marina Clare; http://lattes.cnpq.br/1921551651088660; Vinaud, Marina Clare; Costa, Tatiane Luiza; Castro, Ana Maria de
Taenia crassiceps cysticerci possess antigenic similarities to T. solium cysticerci and because of this and added to the relative easiness in its maintainance in laboratory they are used as an experimental model to T. solium studies. The objective of this study was to analyze the in vitro influence of glucose and insulin on the energetic and respiratory metabolism of T. crassiceps cysticerci exposed to low dosages of anti-helminthic drugs, albendazol and praziquantel, and also to different concentrations of glucose and insulin. Glucose is the main energy source to these parasites in their larval and adult stages and glycogen is their main energetic reserve. Glycogen may be converted into glucose in case of low glucose uptake and is stored in the cysticerci tegument. The cysticerci were collected from the peritoneal cavity of BALB/c female mice experimentally infected and maintained in the animal facilities of IPTSP/UFG. After 24hours of culture in RPMI culture medium supplemented and with glucose, glargine insulin (LANTUS®, Sanofi Aventis), albendazol and praziquantel, the cysticerci were removed and the medium was frozen in liquid nitrogen as to allow the metabolix stasis. Afterwards this culture medium was analyzes through HPLC as to permit the quantification of the following organic acids related to the carbohydrates metabolism: lactate and pyruvate, intermediary metabolism: citrate, α-ketoglutarate, succinate, fumarate, malate and oxaloacetate, fatty acids metabolism: β-hydroxibutyrate and propionate. It was possible to detect propionate and β-hydroxibutyrate secreted by T. crassiceps cysticerci which indicates the fatty acids oxidation as an alternative energy xv source used by the initial stage cysticerci which are in rapid growth. Also the detection of organic acids from the citric acid cycle indicates the in vitro aerobiosis performed by the initial stage cysticerci.
Staphylococcus aureus em tonsilas de pacientes com faringotonsilite aguda recorrente: prevalência, perfil de suscetibilidade e caracterização genotípica
(Universidade Federal de Goiás, 2013-10-17) Cavalcanti, Veraluce Paolini; Braga, Carla Afonso da Silva Bitencourt; http://lattes.cnpq.br/7673897995590123; André, Maria Cláudia Dantas Porfirio Borges; http://lattes.cnpq.br/1475834090578722; André, Maria Cláudia Dantas Porfirio Borges; Cardoso, Juliana Lamaro; Cardoso, Alessandra Marques
The bacterial pharyngotonsillitis are infections of the upper airways that occurs predominantly in children and adolescents. Due to the composition of the oral microbiota is difficult to clarify the role of each organism in the etiology of the disease. The presence of bacteria that produce beta-lactamase interferes with the effectiveness of beta-lactam antibiotics, the most commonly drug used in treatment of these infections, promoting the recurrence of the disease. S. aureus is one of the most common pathogen in the etiology of tonsillitis and its relevance is due to the ability of antimicrobial resistance and persistence in the tissues of the tonsils. Tonsillectomy is indicated in cases of recurrent tonsillitis after several failures in the antibiotic terapy. The aim of this study was to determine the prevalence of S. aureus in the tonsils of patients undergoing tonsillectomy, in a teaching hospital in Goiânia, the antimicrobial susceptibility profile and the genetic characterization of isolates. Tonsils obtained from 123 patients were processed, the microorganisms identified and submitted to antibiogram by conventional techniques. The isolates that presented cefoxitin resistance were submitted to tests to determine the minimum inhibitory concentration - MIC for oxacillin and to detect the presence of the mecA gene. All isolates were subjected to PCR for detection of Panton-Valentine leukocidin gene and to PFGE to determine the genetic similarity among them. It was identified 60 S. aureus isolates from 49 patients (39.8%). There were no significant difference in prevalence by sex and, the average age of male patients was lower (8.2 years) (p<0.001) than the female patients (15.3 years). Nine out 49 patients(18.4%) presented two or more different S. aureus isolates. The isolates presented resistance of 85.0%, 10.0%, 15.0%, 3.3%, 10.0%, 3.3%, 18.3% and 8.3% to penicillin, amoxicillin + ácido clavulânico, cefoxitin, ceftriaxone, erythromycin, trimethoprim/sulfamethoxazole, ciprofloxacin and tetracycline, respectively. All isolates were sensitive to linezolid and rifampin. Six erythromycin-resistant isolates (10.0%) showed inducible resistance (MLSbi) to clindamycin and quinupristin/dalfopristin. Eight isolates (13.3%) were resistant to three or more classes of antimicrobials. Despite the resistance to cefoxitin be considered a marker of the presence of the mecA gene only in two resistant isolates it has been found, xv suggesting that the cefoxitin resistance should be mediated by other mechanisms, such as the overproduction of beta-lactamases. None of these isolates showed resistance to more than two classes of antimicrobials. Among the sixty S. aureus isolates, only carried the gene encoding the Panton-Valentine leukocidin. This isolate presented resistance to five classes of antimicrobials and phenotype D. PFGE analysis grouped 36 (60,0%) of 60 isolates in 10 clusters (>80% similarity), since no one specific clone was associated with colonization of the tonsils. It was observed different patients carrying S. aureus isolates genetically identical or with a high level of similarity (>80%), suggesting in these cases, a common origin. The high prevalence of S. aureus in tonsils suggests an ability to colonize the surface and/or the persistence in the tissues of the tonsils. The isolation of MDR bacteria can promote cross-resistance to other bacteria commonly associated with recurrent tonsillitis. The results point to change the paradigm of diagnosis and treatment of recurrent tonsillitis in order to enable the correct use of antimicrobials to reduce the recurrence which is the main cause of tonsillectomy.
Estudo prospectivo de infecção por calicivírus (norovírus e sapovírus) em pacientes submetidos a transplante alogênico de células progenitoras hematopoiéticas
(Universidade Federal de Goiás, 2013-12-20) Lemes, Lucianna Gonçalves Nepomuceno; Souza, Menira Borges de Lima Dias e; http://lattes.cnpq.br/0054562567103606; Souza, Menira Borges de Lima Dias e; Fiaccadori, Fabíola Souza; Leite, José Paulo Gagliardi
The calicivirus (norovirus and sapovirus) are important etiologic agents of acute gastroenteritis. Recent studies show that in immunocompromised patients such as those undergoing allogeneic hematopoietic stem cell transplantation (HSCT), norovirus infection can lead to worsening of symptoms and be confused with clinical symptoms of graft versus host disease (GVHD). However, calicivirus screening is not performed, routinely, as part of the patients’ follow-up laboratory exams. The main objective of this study was to evaluate the occurrence of norovirus (NoV) and sapovirus (SaV) in patients who underwent HSCT, and to conduct the molecular characterization of the samples positive for these viruses. Fecal samples were collected weekly, and serum samples were obtained every two weeks of ten patients who underwent HSCT, for a minimum period of five months and a maximum of one year. The secretor status was determined by an enzyme immunoassay and the detection of calicivirus was performed by RT-PCR using primers specific for a partial region of the gene encoding the NoV genogroup I and II (GI and GII) and SaV capsid protein. The genomic sequencing was performed for positive samples. The results showed that from ten patients participating in the study, eight had diarrhea. Among these, six (60%) had positive samples for NoV, and all of them had a secretor phenotype. The duration of NoV excretion in feces ranged from five to 143 days. Viral RNA was also detected in serum specimens, ranging from 29 to 36 days in the five patients infected with NoV. Three of the six patients had acute intestinal GVHD. Through genomic sequencing and phylogenetic analysis all NoV-positive samples were characterized as genotype GI.3, and because they had a high nucleotide identity, they were all characterized as a single haplotype. The data highlight the urgent need of the inclusion of calicivirus screening in the routine testing performed before transplantation and during follow-up of these patients. This is the first report of the occurrence of NoV in patients undergoing HSCT in Brazil.
Identificação e caracterização das proteínas da parede celular em Paracoccidioides sp.
(Universidade Federal de Goiás, 2014-02-24) Araújo, Danielle Silva; Parente, Ana Flávia Alves; http://lattes.cnpq.br/8717846564577423; Soares, Célia Maria de Almeida; http://lattes.cnpq.br/8539946335852637; Soares, Célia Maria de Almeida; Bailão, Alexandre Melo; Baeza, Lilian Cristiane
Paracoccidioidomycosis (PCM) is the important systemic mycosis in Latin America, with high incidence in Brazil, Argentina, Colombia and Venezuela. The disease has been attributed to the thermodimorphic fungus Paracoccidioides sp.. During the infective process, we can highlight the role of the cell wall, which is a dynamic structure, vital for growth, survival and morphogenesis of the fungus. In addition, this structure is constantly changing in response to environmental signals and different stages of the cycle of the fungus. The interest in the fungal cell wall occurs primarily by lack of this structure in mammalian cells; for this reason cell wall components are promising targets for antifungal drug design. In order to describe the profile of cell wall proteins (CWPs) of Paracoccidioides sp., it was used a proteomic approach coupling nanoscale liquid chromatography to multiplexed mass spectrometry (nanoUPLC-MSE) to identify the CWPs isolated from Paracoccidioides sp. yeast cells and mycelia. Among the identified proteins, it was found a transglycosylase orthologue to the Crh1p, which is a GPI anchored protein, known to be involved in attaching chitin to β-glucan. Adhesins previously described in Paracoccidioides sp. such as enolase, glyceraldehyde-3-phosphate, alcohol dehydrogenase, fructose-1,6-biphosphate aldose and some chaperones were also identified. Moreover, we identified formamidase that was previously described as localized in the fungus cell wall and may be involved in nitrogen metabolism besides contributing with antigenic properties.
Leishmaniose Tegumentar Americana na região Centro-Oeste: avaliação de dados clínicos, epidemiológicos, laboratoriais e moleculares
(Universidade Federal de Goiás, 2014-04-30) Balian, Rosana Pereira Morais; Dorta, Miriam Leandro; http://lattes.cnpq.br/3933395097851681; Dorta, Miriam Leandro; Moraes, Sandra do Lago; Loyola, Patrícia Resende Alo Nagib
Protozoa of the genus Leishmania, which affects the skin and/or mucous membranes, cause American Cutaneous Leishmaniasis (ACL). It is an endemic zoonosis whose numbers of cases in the Brazil Midwest region are growing. In the Goiás state, 2798 ACL cases were been reported between 2007 and 2013. In the present study, our proposal was to investigate the epidemiological and molecular characteristics of ACL patients attended at the ambulatory of the Hospital Anuar Auad in the period 2000 at 2006 and identify the species of Leishmania sp. The study included 152 patients with ACL, 124 were from of the state of Goiás and 28 at Mato Grosso, aged between 6-79 years, and center and thirteen these individuals were male. For diagnosis, clinical, epidemiological and laboratory data were collected, such as direct examination (ED), histopathology (AH), Montenegro skin test (MST), indirect immunofluorescence (IIF). ELISA using crude extract of L. (Viannia) braziliensis was performing. Characterization of Leishmania species was carried out by polymerase chain reaction (PCR). The positivity of ED, AH, MST, IIF and ELISA was 70.6%, 80.9%, 68.9%, 44.2% and 73.0%, respectively. Specific IgG to L. (V.) braziliensis were detecting in 84.7% of patients with mucosal leishmaniasis (ML), significantly higher than those found in patients with cutaneous leishmaniasis (CL), which was 69.0% (p < 0.05). Detection of IgG before and after treatment was performed using ELISA and was observed a statistically significant difference only in samples obtained from patients with CL after 6 and 18 months of treatment. The data serological monitoring of patients with ACL before and after treatment indicates that the total IgG levels tend to decrease after 6 months of treatment. PCR was performed on 69 samples obtained from scraping the edge of lesions of patients with ATL, these, 62 (89.8%) were positive, 53 patients with LC and 9 patients with ML. The samples were characterized by PCR as L. (V.) braziliensis (93.5%) and L. (L.) amazonensis (6.5%). It was found that 25.8% of cases of leishmaniasis in the state of Goiás occurred in Campestre, Goiânia, Aparecida de Goiânia and Niquelândia. By analyzing the frequency of cases of ATL over the months during the years analyzed, it was observed that there was a distribution of ACL in all seasons from 2000 to 2006, with a larger number of cases in winter. PCR represents important and powerful tool in the diagnosis and species identification in ACL in endemic areas. ACL controlling in endemic areas is difficult and requires an accurate idea of its epidemiology.
Identificação das espécies de Candida, avaliação da ultraestrutura e perfil de suscetibilidade de biofilmes aos antifúngicos
(Universidade Federal de Goiás, 2014-05-08) Marreto, Lais Carneiro Naziasene Lima; Costa, Carolina Rodrigues; http://lattes.cnpq.br/2503738140054176; Costa, Carolina Rodrigues; Silva, Maria do Rosário Rodrigues da; Jesuino, Rosália Santos Amorim
The increase in the number of fungal infections by Candida species, as a result of the significant increase of immunocompromised patients emphasizes the importance of studies associated to this genre. The resistance of these strains to antifungal drugs is observe in specific isolates, as well as infections associated with biofilm formation (BF). The aim of this study was to compare the susceptibility profile of sessile and planktonic cells of different Candida isolates, differentiating species by multiplex qPCR and scanning electron microscopy. The sample consisted of 10 C. albicans, 10 C. parapsilosis and 1 C. tropicalis. The genetic material was extracted by phenolchloroform extraction of the sample C. tropicalis, ATCC 28367 (C. albicans) and ATCC 22019 (C. parapsilosis) for use in a species-specific multiplex qPCR assay with analysis of differences in the curve fusion. Microdilution plate test and reduction of the tetrazolium salt for 21 isolates were performed for the susceptibility of sessile and free cells, respectively. The evaluation of the ultrastructure of the biofilm was performed on strips of polyurethane with visualization by scanning electron microscopy. The amplified products from C. albicans obtained three melting peaks with values of 75,2 °C, 76,8 °C and 79,8 °C, while C. tropicalis and C. parapsilosis showed one peak with values 73,5 °C and 78,5 °C, respectively. The identification test showed to be reliable for the identification of the most common species in the study. All of Candida yeasts were able to form biofilm with own organization of each specie. The in vitro susceptibility test of planktonic cells demonstrated that amphotericin B and caspofungin showed better activity compared to fluconazole and voriconazole who obtained 19% of resistant strains, all C. albicans. In assessing of the CIMs50% and CIMs80% of sessile cells, we observed an increase of at least 60 times the values of planktonic cells. It was observed a high percentage of BF that demonstrates the need for diagnosis and combat against this biomass formed by Candida species. The BF is a virulence factor determinant for decrease of susceptibility to antifungal agents thus compromising antifungal therapy.
Avaliação da atividade antifúngica e mecanismo de ação de compostos naturais e sintéticos em leveduras do complexo Cryptococcus neoformans
(Universidade Federal de Goiás, 2014-05-14) Silva, Kamila Pereira da; Fernandes, Orionalda Fatima Lisboa; http://lattes.cnpq.br/9826992750728262; Silva, Maria do Rosário Rodrigues; Paula, José Realino de
Fungal infections caused by yeasts of Cryptococcus neoformans complex have increased considerably among immunosuppressed especially in patients infected with human immunodeficiency virus (HIV). Antifungal drugs available to treat these infections have a wide spectrum of action, but the high costs, side effects, besides the acquisition of resistance of fungi make their limited effectiveness. In these circumstances, the search for new drugs is needed. In the present study, we evaluated the antifungal activity of tamoxifen, bisabolol, indinavir, didanosine, UFMG, Labiocon 241, solasodine, Labiocon 237, Clonazepam, semicarbonado Benzaldehyde, Cardanol, AB 36 and three natural extracts of Psychotria spp, Sclerolobium paniculatum, Hymenaea courbaril and cell viability and mechanism of action of the compound showed antifungal activity of 20 isolates of Cryptococcus neoformans. To evaluate the antifungal activity , we used the in vitro susceptibility to yeast broth microdilution protocol M27 - A3 ( CLSI 2008) and to verify the mechanism of action, flow cytometry was performed with the cellular marker Propidium Iodide ( PI ) and cell viability assay with MTT salt. Among the compounds analyzed, only tamoxifen and olanzapine showed inhibitory action on fungal samples at concentrations 128-256 μg/mL. Analysis by flow cytometry was performed with tamoxifen and showed that the compound did not alter the cell membrane of Cryptococcus neoformans ATCC 28957 , however, the cell viability test showed that tamoxifen was able to inhibit the metabolism of the fungus in different concentrations ranging 4-1024 μg/mL.
Identificação e caracterização molecular do vírus dengue em indivíduos sintomáticos atendidos na rede pública de saúde de Goiânia – Goiás, durante o período epidêmico 2012-2013
(Universidade Federal de Goiás, 2014-07-18) Guimarães, Vanessa Neiva; Fiaccadori, Fabíola Souza; http://lattes.cnpq.br/0993842409303174; Souza, Menira Borges de Lima Dias e; Cardoso, Divina das Dores de Paula; Fiaccadori, Fabíola Souza
Dengue is a major challenge to public health in Brazil and worldwide. Dengue virus (DENV) is an arbovirus of the family Flaviviridae, genus Flavivirus, classified into four antigenically distinct serotypes DENV1-4. Since the introduction of DENV1 in Goiás, the current situation consisted of successive epidemics of dengue fever with change among prevalence serotypes. One of the factors that have been associated with the pathogenesis of this infection is the occurrence of a cross-serotype immune response in secondary infections. Thus, the identification of circulating serotypes and also the characterization of the genomic variants have been considered important on disease surveillance, since the introduction of new variants can be a risk factor for severe dengue. In this context, the present study aimed to investigate the occurrence of DENV infection in individuals treated at health centers in Goiânia, Goiás in the epidemic period 2012-2013, using serological and molecular methods for determining the rate of positivity and identification of serotypes/genotypes in the circulating virus. For this, 278 samples from suspected individuals of DENV infection, which were submitted to the research of serological markers NS1, IgM and IgG using a commercial kit, as well as viral RNA detection and serotype identification by RT-PCR for region C-prM. The positivity for infection was 43.9%, with rates of 30.5%, 13.6% and 20.5% for NS1, IgM and RNA, respectively. Considering the RNA-positive samples, 31.5% were DENV1 and 68,5% were DENV4, which were subjected to nucleotide sequencing of the E/NS1 junction to characterize the viral genotype. Phylogenetic analysis classified the DENV1 samples as genotype V, that differentiated into two clades, which may reflect the occurrence of genetic variants with differences in evolutionary history. For DENV4 samples, classified as genotype II, was observed a phylogenetic proximity between the strains analyzed and others strains of the country and Latin America. These results demonstrate the effectiveness of the combination methods in the diagnosis of DENV infection. Still, the molecular evaluation has proven crucial to establish a complete data base to be used for surveillance studies, collaborating with investigations of the dynamics of the virus and the expected profile for future epidemics.
Avaliação da ação da IL-17 em macrófagos peritoneais murinos
(Universidade Federal de Goiás, 2014-07-25) Martins, Lohane Suzart; Oliveira, Milton Adriano Pelli de; http://lattes.cnpq.br/2152513705182408; Oliveira, Milton Adriano Pelli de; http://lattes.cnpq.br/2152513705182408; Galdino Júnior, Hélio; Abrahamsohn, Ises de Almeida
L. (L.) amazonensis is one of the species that cause American tegumentary leishmaniasis. Resistance to Leishmania infection is associated with Th1 lymphocytes, which produce IFN-γ and activate macrophages and promote the death of the parasites. However, during the infection, other T cells subtypes can develop, such as the Th17 subtype, which produce IL-17. The role of IL-17 is known in inflammatory diseases and infections with extracellular pathogens, and its function is associated to the migration and activation of neutrophils. In Leishmania infection, it is not clear whether this cytokine would influence the activation of a phenotype that controls infection. Aim: To assess whether IL-17 interferes with activation of murine macrophages and with their Leishmania amazonensis leishmanicidal. Methods: Thyoglycolate-elicited peritoneal macrophages from BALB/c or C57BL/6 mice were primed with IL-17, IFN-γ or IL-4, activated or not with lipopolysaccharide (LPS) and infected with L. (L.) amazonensis promastigotes. Culture supernatants were harvested 48 h after the stimulus to assess nitric oxide (NO) and IL-12p40 production. The cells were used to determine the phagocytic, leishmanicidal and arginase activity. Results: Macrophages from BALB/c and C57BL/6 mice stimulated with IL-17 and LPS produced IL-10 and increased arginase activity. IL-17 was not able to induce NO production by these cells, but promoted a small increase in IL-12p40 secretion by C57BL/6 mice macrophages. IL-17 favored the proliferation of Leishmania in macrophages from BALB/c mice, but not in C57BL/6 mice. A synergism was observed between IL-17 and IL-4 that increased IL-10 production and arginase activity in macrophages from BALB/c mice. In macrophages from C57BL/6 mice, however, a synergism was observed between IL-17 and IFN-γ in the stimulation of NO production. Conclusion: These results suggest that IL-17 may participate in the activation of macrophages toward either phenotype depending on the mouse strain. The finding that IL-17 impairs the control of L. (L.) amazonensis in BALB/c mice was not observed in C57BL/6 mice. IL-10 production in macrophages stimulated with IL-17 suggests a regulatory role of this last cytokine.
Avaliação histopatológica de lesões por queimaduras de 3°grau em ratos Wistar com e sem diabetes, tratados com o laser de baixa potência
(Universidade Federal de Goiás, 2014-07-25) Mendonça, Diego Eterno de Oliveira; Loyola, Patrícia Resende Alo Nagib; Lino Júnior, Ruy de Souza; http://lattes.cnpq.br/0372118837748010; Lino Júnior, Ruy de Souza; Loyola, Patrícia Resende Alo Nagib; Parente, Leila Maria Leal; Moraes, Julia de Miranda
The use of low level lasers in the burning wounds healing has been described in the literature with a great variety of treatment parameters such as multiplicity of wave lenghts, energetic dosages and time of use. Objective: the aim of this study was to compare the effect of low potency AsGaAl laser in the third degree burning wounds healing in Wistar rats with and without diabets. Methodology: this project was approved by the ethics committee in Animals use from UFG, protocol number 007/2012.100. The animals were distributed into 4 groups: a) control group non diabetic; b) treated group non diabetic; c) diabetic control group; d) diabetic treated group. The treated groups received an energetic dosage of 3 J/cm2 in the first 7 days and 6 J/cm2 after the seventh experimental day. The animals received changes of the occlusive bandages embedded with silver sulphadiazine and all of them were accompanied throughout 3, 7, 14, 21 and 30 days for the macroscopic, morphometric and microscopic aspects evaluation. The parametric "t test" was used for the statistical analysis. Results: the comparison of the wounds from the diabetics and non diabetics animals showed that the treatment with low laser induced a significant reduction (p<0.05) of the contraction area; macroscopically there were no differences between groups. Regarding the microscopic findings there was a significant increase in angiogenesis (p<0.05) and increase in the fibrin presence (p<0.05) when compared the treated groups from diabetic and non diabetic animals in 3rd and 14th days of experiment. In the treated diabetic animals the angiogenesis in the 14th day and the number of fibroblasts in the 14th and 21st days of experiment presented a significant difference (p<0.05) when compared to the non treated group. Also there was greater collagen disposition throughout the 14th, 21st and 30th days of experiment in all treated groups when compared to the control groups. In conclusion the low laser treatment reduced the injury in long term, stimulating the angiogenesis, fibrin deposition, fibroblasts and collagen deposition.
Caracterização proteômica da fração nuclear da forma leveduriforme de espécies de Paracoccidioides
(Universidade Federal de Goiás, 2014-08-01) Oliveira, Lucas Nojosa; Casaletti, Luciana; Soares, Célia Maria de Almeida; http://lattes.cnpq.br/8539946335852637; Soares, Célia Maria de Almeida; Borges, Clayton Luiz; Pereira, Maristela
Paracoccidioidomycosis (PCM) is an endemic disease in Latin America caused by fungi of the genus Paracoccidioides which is composed of two species P. lutzii e P. brasiliensis. The fungus grows as mycelium in saprophytic environment, and as yeast in host. Paracoccidioides spp present multinucleated yeast cells. The characterization of the nuclear proteome an important step to understand the biology of the fungus, and contributes to future studies of possible pharmacological targets. In this study we aimed to characterize the profile of the nuclear proteome of Paracoccidioides species in yeast form. A total of 570 proteins were identified and classified as nuclear, using a combination of method sample enrichment, proteomic of high accuracy technique, NanoUPLC-MS E , and bioinformatic filters. The results revealed important proteins related to DNA maintenance, regulation and gene expression, synthesis and processing of messenger and ribosomal RNA and nuclear-cytoplasmic traffic. Uncharacterized proteins and proteins commonly known as cytoplasmic were identified in fraction enriched nuclear and require further studies to demonstrate its nuclear roles. This is the first descriptive nuclear proteome of Paracoccidioides spp. and opens the way for new researches.
Análises proteômicas de conídios do fungo patogênico humano Paracoccidioides sp
(Universidade Federal de Goiás, 2014-08-22) Moreira, André Luís Elias; Bailão, Alexandre Melo; http://lattes.cnpq.br/5415221996976886; Borges, Clayton Luiz; http://lattes.cnpq.br/8867708267053410; Borges, Clayton Luiz; Bailão, Alexandre Melo; Parente, Ana Flávia Alves; Lima, Patrícia de Souza; Brito, Wesley de Almeida
The paracoccidioidomycosis (PCM) is a systemic disease caused by the thermo dimorphic fungus Paracoccidioides spp.. The route of infection of the PCM occurs by the inhalation of conidia or mycelia fragments. Until now, no proteomic studies were performed with conidia of Paracoccidioides sp. In this sense, characterize the proteome of the conidia, may contribute to the detailed knowledge of the proteins expressed during the propagation phase and their potential roles in virulence and pathogenicity, providing possible targets for antifungal strategies. For the conidia production, the mycelia of isolate Pb01 (ATCC MYA-826) were cultured in potato agar medium during 90 days at 18 ºC. After production, the conidias were collected and purified. The proteins were extracted and subjected to tryptic digestion with subsequent identification by NanoUPLC-MSE. We identified a total of 242 proteins of conidia of Paracoccidioides. The in silico analysis were used to characterize the proteins present in Paracoccidioides conidia. Proteins as GAPDH, enolase, triosephosphate isomerase, and some glycoproteins like ECM33 and β-1,3-glucanosyltransferase gel2 were identified during analysis. Some of these have been described as adhesins in Paracoccidioides and in other pathogenic fungi. Were also identified proteins related to signal transduction pathways, such as: Ras GTPase, RhoA GTPase and calmodulin, described in other fungi involved in morphologic changes. Proteins related to evasion, defense and virulence of the fungus, such as HSP90, catalase, mitochondrial peroxiredoxin Prx1, have been described with functions related to temperature shifts or oxidative stress provided by the host environment, were also identified. These results highlight that Paracoccidioides conidia contain proteins that can contribute to its maintenance in the environment and have molecules related to important processes necessary for the initial steps of infection in the host.
Efeitos do laser de baixa potência na leishmaniose experimental: avaliação sobre a lesão cutânea (in vivo) e efeitos sobre leishmânia (in vitro)
(Universidade Federal de Goiás, 2014-09-01) Rocha, Jhonathan Gonçalves da; Loyola, Patricia Resende Alo Nagib; http://lattes.cnpq.br/7172668220681444; Loyola, Patrícia Resende Alo Nagib; Celes, Mara Rúbia Nunes; Avelar, Juliana Boaventura
Leishmaniasis is neglected infectious and parasitic diseases caused by protozoa of the genus Leishmania, having different clinical forms according to the type of host-parasite relationship established, the main ones being: Leishmaniasis Visceral and Cutaneous Leishmaniasis, object of this study. The fact that conventional treatments present limitations and difficulties for patients coupled with increasing use of low level laser therapy in the treatment of skin lesions of various etiologies, specifically with Low power laser, led us to evaluate the possible consequences of such treatment in American cutaneous leishmaniasis in both murine experimental models (in vivo), with the aim of evaluating the dynamics of healing of ulcerated lesions, as in cultured parasites (in vitro), with the use of techniques that allowed us to evaluate the interference of therapy in cultures of the parasites isolated from the MAB-6 Leishmania (Leishmania) amazonensis Leishmanias belonging to the Bank of the Midwest, with regard to the ability to multiply, viability, integrity and infective capacity. The tests "in vivo" concluded that the use of low-power laser is an interesting tool in the treatment of ulcerative lesions caused by cutaneous leishmaniasis since it leads to decreased edema, stimulates microbicidal action of macrophages and promotes the maturation of collagen. Already in essays "in vitro", it was found that the action of the treatment leads to a static effect on the growth curve of the parasites. The viability tests showed a small increase in the number of parasites killed after treatment and reduction in mitochondrial activity. The low level laser therapy proved incapable of promoting the apoptotic process and modify the composition of the plasma membrane of the parasite, which suggests that the laser treatment did not affect the ability of infection. Instead, there was a rise in titers of viable parasites present in the paws of mice infected with Leishmania treated.
Caracterização de uma nova linhagem de célula dendrítica: AP284
(Universidade Federal de Goiás, 2014-10-23) Oliveira, Pollyana Guimarães de; Oliveira, Milton Adriano Pelli de; http://lattes.cnpq.br/2152513705182408; Oliveira, Milton Adriano Pelli de; http://lattes.cnpq.br/2152513705182408; Silva, Juliana Reis Machado e; Shio , Marina Tiemi
Dendritic cells (DCs) are a heterogeneous group of cells and the major activators of naive T lymphocytes. During antigen presentation, different types of cytokines are produced and they will interfere with the immune response profile generated. IL-12p70 promotes the differentiation of Th1 phenotype and IL-23 p enhances the Th17 response. Given the major role of IL-12 and IL-23 on the stabilization of the acquired immune response, the production of these cytokines is highly controlled and it is an issue for several studies. The objective of this work was to characterize a new lineage of DC, described as AP284, based on their surface markers and evaluate the ability of these cells to produce IL-12p40, IL-12p70 and IL-23 after different stimulus in vitro. Additionally it will be evaluate their phagocytic capacity and their ability to induce a Th1 and Th17 response in vivo. The expression of the markers was analyzed by flow cytometry and the quantitation of IL-12 and IL23 was performed by ELISA after stimulation of cells AP284 in vitro with lipopolysaccharide (LPS) and Escherichia coli. AP284 cells express MHC class II, CD11c and 33D1 on its surface, which are characteristic markers of DCs. After stimulus with LPS or E.coli, AP284 cells produce large amounts of IL-12p40 and IL-23 but do not produce IL-12p70. The production of IL-12p70 was not detected even when IFN-γ is added to prime cultures, moreover, priming the cells with IFN-γ promoted inhibition of IL-12p40 and IL-23. Our data sugest that the AP284 is a DC17 lineage. Thus, AP284 can be an important tool to study the mechanisms of induction or regulation of IL-23 production and a possible tool to study generation and maintenance of a Th17 profile.
Atividade larvicida da Persea americana (Lauracea) sobre Aedes aegypti (Diptera, Culicidae) evidenciada por modificações morfohistológicas
(Universidade Federal de Goiás, 2014-11-03) Guimarães, Antonella Del Buono; Silva, Ionizete Garcia da; http://lattes.cnpq.br/5021551669347602; Arruda, Walquíria; Silva, Heloísa Helena Garcia da; Silva, Ionizete Garcia da
Dengue has impact on public health of the intertropical regions of the world and its main vector Aedes aegypti. The measures most commonly used to control this vector, so far, are the synthetic insecticides, which applied on a large scale for a long period, did show resistance. Thus, chemical compounds from plants can be substitutes for synthetic insecticides, with the advantages of resistance retardation, due to the complexity of its components, easy degradation, lower toxicity to man and to be a safer alternative to the environment . Assays crude ethanol extract (cee) of Persea americana stem bark were performed in polystyrene cups, solutions containing 100 ppm of P. americana cee, previously dissolved in dimethylsulfoxide (DMSO) and water public Goiânia system. In each glass were placed 5 larvae of 3rd stage (L3) of the mosquito Aedes aegypti. The study was made of morfohistológic modifications from observations made with 4, 8, 12, 16, 20, 24 and 48 h treatment with the cee. In each period, the larvae were collected, fixed, dehydrated, infiltrated and embedded in historesin. of 3μm thick cuts were made in microtome semiautomatic, stained with hematoxylin / eosin (HE), analyzed and photomicrographed under a light microscope. For scanning electron microscopy (MEV) the treated and control larvae were fixed, dehydrated, dried in a critical point apparatus, metallized and analyzed in electron microscope. The morfohistológic changes were observed from 24 hours after treatment of larvae L3 with P. americana stem bark cee. The analysis of histological sections of the midgut of Ae. aegypti in light microscopy showed the presence of vacuoles in the cytoplasm of epithelial cells, large amount of secretion and folding of membrane peritrophic larvae treated for 48 hours. There was extrusion whole food content together with peritrophic matrix, thus suggesting the P.americana. toxic action on the gut of the larvae. These changes did not occur in the control group. There was no evidence of external changes by MEV.
Avaliação de diferentes lotes de soro bovino fetal no preparo de meio para cultura de Leishmania (Viannia) braziliensis
(Universidade Federal de Goiás, 2015-02-27) Santos, Jéssica Cristina dos; Oliveira, Milton Adriano Pelli de; http://lattes.cnpq.br/2152513705182408; Castro, Ana Maria de; Pinge Filho, Phileno; Oliveira, Milton Adriano Pelli de
Promastigote forms of Leishmania (V.) braziliensis can be mantained in vitro for use in research. The bovine fetal serum (BFS) is an important component in several culture medium, however, its composition varies from batch to batch. This work aims to evaluate the performance of different BFS batches for preparation of Leishmania (V.) braziliensis culture media, comparing information supplied by different known techniques. PPS6m and CSA7c leishmanias strains were from samples obtained from American Tegumentary Leishmaniasis (ATL) patients and five different batches of BFS were tested in Grace’s medium. The parasite growth curve was evaluated by a daily hemocitometer counting, the metabolism was evaluated by the parasite’s ability to metabolize 3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT), the percentage of promastigote metacyclic forms was quantified by an agglutination test using Bauhinia purpuera (BPL) lectin and the ability to promote mice lesions evaluated by a weekly measurement of lesions. The biochemical components of the BFS batches was also performed. It was observed that two or more passages in culture were necessary to identify bad batches of BFS by the growth curve analysis, while the metabolic analysis showed that an inadequate BFS batch of provided a metabolic rate higher or equal to the best BFS batch. A lower percentage of metaciclic forms of PPS6m strain was observed in one of the media, however, the percentage of metacyclic promastigotes was similar among all the CSA7c culture media, independently of the parasite number. Besides, the stationary phase parasite’s ability to cause lesions in BALB/c mice did not depend on the SBF batch present in the cultures. Data reveals that the best method to select good culture serum is to observe the growth curve for two or more passages, besides this, bad SBF batches are able to produce parasites with the same ability to infect mice.
Análise bioquímica da niclosamida no metabolismo de cisticercos de Taenia crassiceps
(Universidade Federal de Goiás, 2015-03-09) Costa, Marco Vítor Silva de Melo; Costa, Tatiane Luiza da; http://lattes.cnpq.br/8219041935358486; Vinaud, Marina Clare; http://lattes.cnpq.br/1921551651088660; Vinaud, Marina Clare; Silva, Luciana Damacena; Bezerra, José Clecildo Barreto
Experimental studies with Taenia crassiceps have been used to demonstrate its metabolic alterations in biochemical pathways (energetic and respiratory) in response to the presence of drugs. The use of T. crassiceps as an experimental model offers conditions to reproduce the neurocysticercosis infection one of the most severe form of human cysticercosis. This study aimed the analysis of the in vitro influence of niclosamide, an antihelminthic drug, on the metabolism of carbohydrates and fatty acids of T. crassiceps cysticerci (ORF strain). 20 larval stage cysticerci were cultured into 5mL of RPMI (Gibco) supplemented culture media, in 6 well culture plates, exposed or not to niclosamide (1, 2, and 3 uM). The control groups were performed with cysticerci not exposed to the drug, exposed to ethanol in the concentration used to dissolve the drug. The cysticerci were cultured at 37ºC for 24 hours. After this period the cysticerci were removed from the culture medium and both were frozen with liquid nitrogen. Afterward, the samples were processed for the HPLC analysis. Accordingly to the mode of action of this drug, which is to interfere in the electrons chain transport, the succinate concentrations were altered in the secretion/excretion of this parasite. Therefore, it is possible to conclude that in the concentrations used, this drug caused little alteration in the metabolic pathways of the parasite.
Análise das atividades antimicrobiana e citotóxica de actinobactérias isoladas de diversos habitats
(Universidade Federal de Goiás, 2015-03-18) Oliveira, Bruno Francesco Rodrigues de; Vieira, José Daniel Gonçalves; http://lattes.cnpq.br/1742731776579730; Vieira, José Daniel Gonçalves; Ulhoa, Cirano José; Souza, Keili Maria Cardoso de
The actinobacteria constitute a wide and diverse phylum of gram-positive bacteria with a high content of guanine and cytosine in their DNA, standing out by the production of secondary metabolites with antibiotic activity. The treatment of multi-drug resistant pathogens infections and cancer represents a major challenge for modern medicine and bioactive microbial metabolites constitute a rich source of antimicrobial and antitumor drugs. The main aim of this study was to analyse the potential antimicrobial and cytotoxic activities of actinobactéria isolated from various habitats. The evaluation of these bioactivities was accomplished by the application of a set of qualitative screening in vitro assays. From a total of 19 morphospecies, isolated from the goiano Cerrado soil, 16 were evaluated in the primary antimicrobial activity tests, of which 5 (31,25%) antagonized the growth of indicator microorganisms. The ECL of the morphospecie BC-A22 and the crude extracts of the strain ADU 1.3 stood out for their strong antibiotic action against MRSA. The most of the crude extracts exhibited lytic action on the staphylococcal cell wall. Only ECLs of morphospecies GUARA 1 and PEG 23 and extracts of the strain PEG 30 were cytotoxic against A. salina in low concentration. None of the extracts showed an intercalation effect on the DNA molecule. With exception of BC-A22, the other six isolates were morphologically characterized as members of the Streptomyces genus. The morphospecie ADU 1.3 was molecularly identified as Streptomyces sp., highly likely to consist of a new species. The results of this initial phase of microbial bioprospecting highlight that all the isolates are potential producers of antibiotics biomolecules, excelling the morphospecie BC-A22, which exhibited a strong antimicrobial activity against the clinical isolates of MRSA and most of gram-negative bacteria.
Monitoramento e caracterização molecular de adenovírus humanos em amostras provenientes de pacientes submetidos ao transplante de células progenitoras hematopoiéticas
(Universidade Federal de Goiás, 2015-03-19) Santos, Hugo César Pereira; Souza, Menira Borges de Lima Dias e; http://lattes.cnpq.br/0054562567103606; Souza, Menira Borges de Lima Dias e; Loyola, Patricia Resende Alo Nagib; Costa, Paulo Sérgio Sucasas da
The human adenoviruses (HAdV) infect people of all ages worldwide, causing a wide range of clinical syndromes, depending on the viral type. In immunocompromised hosts, as transplanted patients, HAdV infection can result in a bad prognosis. Some aspects of viral pathogenesis, such as the association between viremia and/or viral load with disseminated disease and the optimal moment to start the therapy, are still not well established in adult patients that have undergone allogeneic hematopoietic stem cell transplantation (ASCT). Therefore, the main objective of this study was to monitor ASCT recipients for HAdV occurrence, and also correlate viral positivity, viral load and molecular variant with clinical symptoms and patients’ prognosis. For this, stool and serum from 21 patients were monitored in a 2 years period (from October/2012 to October/2014). Serum and fecal samples were screened by Nested-PCR, using primers targeting a partial region of the hexon gene (143bp). Fecal samples were further screened by a commercial enzyme immunoassay (EIA). In total, 57% of the patients had at least one positive sample (serum or stool) for HAdV. Patients presented high viral load (varying from 7,7x103 to 2x108 copies/mL), with a higher viral load in stool when compared to serum. Positive samples were submitted to genomic sequencing, revealing the occurrence of HAdV from C, D and F species. The main clinical symptom presented by infected patients was diarrhea, and graft-versus-host disease was the main intercurrence; however it was not possible to directly associate viral positivity to cause of death. We hope to contribute for a better understanding of the HAdV infection pattern in patients submitted to ASCT. Our data highlights the importance of the inclusion of HAdV testing in the routine laboratory exams of this group of patients.
Desenvolvimento de metarhizium anisopliae em ninfas de periplaneta americana durante a invasão
(Universidade Federal de Goiás, 2015-03-27) Machado, João Antônio Rangel; Fernandes, Éverton Kort Kamp; http://lattes.cnpq.br/2135541732341157; Luz, Wolf Christian; http://lattes.cnpq.br/1104009511235835; Luz, Wolf Christian; Xavier, Solange; D’ Alessandro, Walmirton Bezerra
The susceptibility of cockroaches to infection with pathogenic fungi can be associated to the host species and their stage/instar. The high resistance of elder nymphs of Periplaneta americana, a synanthropic species in tropical and subtropical regions, against Metarhizium anisopliae is still poorly understood and can be connected to behavioral, physiological and mechanical mechanisms. Fourth and fifth instar nymphs (N4 or N5), treated topically with conidia, removed mechanically part of the conidia from the cuticle, antennae and legs, immediately after treatment and in the following days. A small number (< 5%) of the conidia on the cuticle germinated in the first 24 hours, and quantitative germination of the remaining conidia reached 80% within the next 7 days. In most of the treated nymphs, the fungus failed to reach the hemolymph during the 10 days of the test, and in the others, the number of hyphal bodies ranged from 500 to 8 x 107 per μL hemolymph. Few nymphs (30%) died within 10 days after inoculation, and M. anispoliae developed in 50% of the cadavers. The results confirmed the high resistance of elder P. americana nymphs to M. anisopliae and highlighted the importance of the early stages of the infection during the contact and insect invasion by the fungus. Subsocial behavior patterns and specific physiological factors in the cuticle, not elucidated in this study, are resistance key issues and need to be considered when developing mycoinsecticides for P. americana and other cockroach control.

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