Doutorado em Medicina Tropical e Saúde Pública (IPTSP)
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Item Atividade larvicida e caracterização molecular dos princípios ativos de Magonia pubescens St.Hil. (Sapindaceae) e de Copaifera reticulata Ducke (Leguminosae), visando ao controle de Aedes aegypti(Universidade Federal de Goiás, 2004-12-02) Silva, Heloisa Helena Garcia da; Silva, lonizete Garcia da; Silva, lonizete Garcia da; Rodrigues Filho, Edson; Santos, Regina Maria Geris dos; Rodrigues, Maria do Rosário; Bezerra, Jose Clecildo BarretoDengue is an acute viral disease of great importance to the public health, and its high incidence in the tropical countries is intimately related to the presence of the main vector, the mosquito Aedes aegypti. Throughout the years, attempts to control the vector have been based on the application of synthetic chemical insecticides, which have already began to produce undesirable effects. The modification of the susceptibility and the emergence of generations resistant mosquitoes besides fast proliferation stimulated studies about the activity of natural products on the larvae of A. aegypti, as an alternative measure for control. In this work, phytochemicals studies were accomplished by larvicidal activity of the plants Magonia pubescens St. Hil. (Sapindaceae) and Copaifera reticulata Ducke (Leguminosae), with the purpose of isolating fractions and/or pure substances with insecticide potential. After collection of peels of the M. pubescens stem and C. reticulata oil-resin in natura, extracts obtained were submitted to bioassays, guided-purification and structural identification. For the larvicidal activity assays, 3rd instar larvae of A. aegypti were used. They were obtained from cyclic colony maintained by ten years, at 28±1°C, 80±5% of relative humidity and 12 h photoperiod. Twenty larvae were used for each concentration and the bioassays were carried out in 5 replicate, in an acclimatized ambient similar to colony growth. Control assays were conducted using the same number of larvae in a dimethylsulphoxide and distilled water solution. The mortality of larvae was measured after 24 and 48 h. Fractions, subfractions and pure substances with larvicidal activity, obtained from those procedures, were monitored chemically through thin layer chromatography and analyzed by 1H nuclear magnetic resonance and 13C, and gas chromatography coupled to mass spectrometry. The identified active substance in the M. pubescens was a tannin (C45H36O18 and molecular mass of 864.77 Da) which presented LC50 of 3.1 ppm; from the C. reticulata the acid 3-acetoxi-labda-8(17),13-dien-15-oic was isolated (C22H34O4, and molecular mass of 362 Da) with LC50 of 0.8 ppm. These two active substances presented lethal concentrations with potential use in the actions to control of the A. aegypti.Item Detecção e genotipagem do papilomavirus humano em amostra do colo do útero de adolescentes do Distrito Sanitário Noroeste do município de Goiânia, Goiás(Universidade Federal de Goiás, 2005-03-31) Daud, Lyana Elias Santos; Alves, Maria de Fátima Costa; http://lattes.cnpq.br/1412047788155346; Alves, Maria de Fátima Costa; Martins, Regina Maria Bringel; Villa, Luísa LinaBackground: Human Papillomavirus (HPV) infection is a strong predictor for cervical cancer development. Some HPV types are considered at high-risk and associated with pre-malignant lesions and cervical cancer. Genital HPV infection is highly prevalent in sexually active young women. Actually PGMY09/11 primer system associated with Restriction Fragment Length Polymorphism (RFLP), hibridization or DNA sequencing is one of the most sensitive methods for detecting and typing HPV-DNA in genital samples. Objectives: To detect and identify HPV-DNA genotypes in genital specimens among sexually active female adolescents from Distrito Sanitário Noroeste, Goiânia. Methods: We performed a cross-sectional study among 432 sexually active female adolescents (15-19 years), randomly selected at Distrito Sanitário Noroeste, Goiânia, and served by the Family Health Program. HPV-DNA was detected by PCR assay, in endocervical samples, using the PGMY09/11 consensus primer set. The presence of amplifiable DNA was assessed with -globin primers (PCO3/PCO4). Amplicons generated with PGMY primers were typed with the line blot assay (PGMY-line blot) and RFLP. Results: HPV-DNA was detected in 121 samples yielding a 28% prevalence (95% CI 23.8- 32.5). All samples yielded a -globin amplimer, confirming the DNA adequacy. Thirty different HPV genotypes were identified, the high risk genotypes frequently found being: HPV-16 (6,7%); followed by HPV-51 (5,1%); HPV-31 (4,6%); HPV-52 (4,2%) and HPV-18 (3,5%). PGMY-line blot test identified 54 cases of infection caused by multiple genotypes. The genotypes 16, 51, 18, 53 and 52 were the most frequently associated with cervical co- infection with multiple HPV types. Conclusions: A high prevalence of HPV-DNA was found and a broad spectrum of HPV genotypes was identified, predominantly high risk HPV genotypes. A high frequency of multiple HPV infections was found in the studied population.Item Estudo epidemiológico e molecular da infecção pelo vírus da hepatite B em Afro-descendentes de comunidade isolada no Estado de Goiás (Kalungas)(Universidade Federal de Goiás, 2007-12-18) MATOS, Márcia Alves Dias de; MARTINS, Regina Maria Bringel; http://lattes.cnpq.br/2582896795892370Hepatitis B virus (HBV) infection occurs throughout the world. In Africa, this infection is highly endemic, with the majority of individuals becoming infected during childhood. Although Brazil has been globally considered a country of HBV intermediate endemicity, variable rates have been found in all five Brazilian regions and even inside the same region. This study aimed to investigate the epidemiological and molecular profile of the HBV infection among the Kalunga population in Goiás, Central Brazil, which is considered the largest Afro-Brazilian isolated community. A total of 878 individuals were interviewed about sociodemographic characteristics, risk factors and HBV vaccination. Blood samples were collected from all participants and serum samples were screened by enzyme-linked immunosorbent assay for the presence of HBsAg, anti-HBc and anti-HBs serological markers. HBsAg-positive samples were submitted to HBeAg and anti-HBe detection. HBsAg and anti-HBc positive samples were tested for HBV DNA detection by polymerase chain reaction and genotyping by subsequent restriction fragment length polymorphism (RFLP) analysis and nucleotide sequencing of preS/S region. The overall prevalence of HBV infection was 35.4% (95% CI: 32.3-38.7). HBsAg carrier rate was 1.8% (95% CI: 1.1- 3.0). Multivariate analysis of risk factors showed that increased age, male gender, illiteracy and history of multiple sexual partners were associated with this infection. Isolated anti-HBs was found in 301 (34.3%) individuals who were immune for hepatitis B. HBV DNA was detected in 75% (12/16) of the HBsAg positive samples, in 100% (2/2) of the HBeAg and in 83.3% (10/12) of the anti-HBe positive samples. An occult HBV infection rate of 1.7% (5/295) was found among anti-HBc positive individuals. All genotyped isolates belonged to genotype A by RFLP analysis. Nucleotide sequencing of preS/S region confirmed the circulation of genotype A (subgenotype Aa) in this community. The epidemiological findings indicate that preventive measures, such as additional health education and HBV vaccination programs, are needed to control HBV infection in this population. In addition, the molecular results suggest the introduction of genotype A, subgenotype Aa in Brazil from Africa during the slave trade.Item Caracterização dos genes de NSP4 e VP6 de amostras de rotavírus do grupo A provenientes de crianças da região Centro-Oeste do Brasil(Universidade Federal de Goiás, 2008-04-28) TAVARES, Talissa de Moraes; BRITO, Wilia Marta Elsner Diederichsen de; http://lattes.cnpq.br/7605775995731168; CARDOSO, Divina das Dôres de Paula; http://lattes.cnpq.br/9770835116155857Group A rotaviruses are the major cause of gastroenteritis in children throughout the world. Epidemiological surveys and molecular analysis of rotavirus strains are required for gastroenteritis control and prevention. Studies using VP6, an important immunogenic structural protein, and NSP4, a transmembrane nonstructural glycoprotein which is critical to rotavirus morphogenesis and pathogenesis, have been performed. In this study, 330 rotavirus-positive fecal samples previously obtained from children with or without diarrhea, between 1987 and 2003, in three cities of Central West Region of Brazil (Goiânia, Brasília and Campo Grande), were characterized for VP6- and NSP4-encoding genes. The VP6 and NSP4 genes were amplified by reverse transcription- polymerase chain reaction followed by sequencing and phylogenetic analysis. Detection rates of 84.8% and 78.5% were observed for VP6 and NSP4 genes, respectively. Two distinct genotypes could be recognized for NSP4 (A and B). It was observed that the G9P[6] samples were associated with genotype A, whereas the G1P[6], G1P[8], G2P[8], G3P[8], G4P[8] and G9P[8] samples were associated with genotype B. The analysis of VP6 gene allowed genogrouping of samples in two clusters, genogroups I and II. The G2P[4], G3P[4] and G9P[6] samples were identified as genogroup I, whereas the G1P[6], G1P[8], G2P[8], G4P[6], G4P[8] and G9P[8] samples were identified as genogroup II. In addition, it was showed that samples identified as VP6 genogroup I were associated with NSP4 genotype A and samples identified as VP6 genogroup II were associated with NSP4 genotype B. This investigation described different genetic groups representing diversity of group A rotavirus samples circulating in the Central West Region of Brazil.Item ANÁLISE ESPACIAL DA INFECÇÃO PELO VÍRUS DO DENGUE NO MUNICÍPIO DE GOIÂNIA.(Universidade Federal de Goiás, 2008-06-02) MACIEL, Ivan José; MARTELLI, Celina Maria Turchi; http://lattes.cnpq.br/5867052489026059Dengue is nowadays considered a growing public health problem worldwide. Several outbreaks of dengue have occurred in Brazil in the last two decades, and the country is now considered an endemic area where risk areas for sylvatic yellow fever also coexist. The current manuscript reviews the main epidemiological features of dengue in the world focusing in the peculiarities of the infection/disease progression in Brazil and, specifically, in Central-West Brazil. Some issues related to the challenge of control in the Central-West region and the opportunities for research are also discussed. In Brazil, the re-introduction of the vector (Aedes aegypti) dates 1976-77. The city of Rio de Janeiro (Southeast Brazil) was considered the starting point of viral dispersion to coastal and inland areas, since the first epidemic (DENV-1) in 1986. Brazil reports approximately 70% of the dengue cases in the Americas with the co-circulation of 3 dengue subtypes (DENV-1; DENV-2 and DENV-3). The disease affects mainly the adult population and the surveillance system has detected an increasing trend to hospitalization, disease severity and incidence in children and adolescents. Approximately 500,000 of dengue cases and 158 deaths were reported in 2007 compared to approximately 300,000 and 77 deaths in the previous year in Brazil. The first epidemic in Goiás State (Central-West Brazil) was reported in 1994. Nowadays the three serotypes co-circulate with high incidence rates and a large outbreak was reported in the city of Campo Grande (Mato Grosso do Sul State) (45,843 registered cases). The recent increase in cases related to sylvatic yellow fever, mainly in Goiás State, represents a public health warning related to vector surveillance and control.Item Caracterização bioquímica da endoxilanase recombinante (HXYN2r) do fungo termofílico Humicola grisea var. thermoidea e sua aplicação na sacarificação de resíduos agrícolas(Universidade Federal de Goiás, 2008-06-30) CARVALHO, Wagner Rodrigues de; FARIA, Fabrícia Paula de; http://lattes.cnpq.br/3739169267521003Xylanases have been used in the biobleaching of paper pulps, in bioconversion of plant biomass, for food and feed industries, among others. For successful selection of xylanases suitable for specific industrial applications it is important to characterize enzymes isolated from different sources. The thermophilic fungus Humicola grisea var. thermoidea is described as a good producer of extracellular endoxylanases and the Hxyn2 gene from this fungus was isolated and expressed in yeast Pichia pastoris. The aim of this project was focused on the production, purification and biochemical characterization of the recombinant HXYN2 (HXYN2r) enzyme and application on enzymatic hydrolysis of lignocellulosics substrates. The culture conditions of P. pastoris in flasks, using the 12.3 transformant and BMMY-U medium, were optimized and the best xylanolytic activity was 478.2 U/mL after 96 h, with a protein concentration of 100 mg/L, using 2.34% (w/v) of nitrogen sources (yeast extract plus urea 1.34:1.0% - w/v), 1% (v/v) of methanol, and OD600 of 10. The HXYN2r enzyme was purified by gel filtration chromatography with a yield of 6.4% and showed optimum pH and temperature values of 6.5 and 60ºC, respectively, maintaining 100% of initial activity after 4 h of incubation at pH 5.5, 6.5 and 7.5. The half-life time was 18 min at 60ºC and the enzyme was 100% stable after 3 h of incubation at 50ºC. Km and Vmax values were 7.9 mg/mL e 235.4 μmol/(mL.min), respectively. The HXYN2r enzyme were used on the enzymatic hydrolysis of sugarcane bagasse (SCB), corn cob (CC) and foliar sample of sugarcane (FSC) alone or with the enzymes of xylanolytic and cellulolytic system produced by H. grisea fungus. For enzymatic hydrolysis experiments, the substrates were milled and pretreated with 0.25% (w/v) of H2SO4 by 30 min. On the enzymatic hydrolysis the best conversion yield of hemicellulosic fractions were obtained using PpHXYN2r supplemented with EHg: 42.8% to CC, 9.6% to SCB and, a mean of 20% to foliar samples.Item Enfermidades infecciosas em comunidade indígena Terena de Mato Grosso do Sul(Universidade Federal de Goiás, 2008-08-02) AGUIAR, José Ivan Albuquerque; NETTO, Joaquim Caetano de Almeida; http://lattes.cnpq.br/3444498706763045The health conditions of Brazilian indigenous population are few recognized with limited information available. A survey was carried out among indians Terena, people that inhabit on the municipalities of Sidrolândia and Dois Irmãos do Buriti, Estado de Mato Grosso do Sul, Brazil. Was studied the prevalence of infection by intestinal parasites, infection markers for viral Hepatitis A, B and C, and for antibody against the viral diseases; Poliomyelitis 1, 2 and 3, Measles, Yellow Fever and Hepatitis B. The results were stratified by age and revealed that the parasitic infection affects 73.5% of the population studied, with high prevalence of the Blastocystis hominis. In the population above 10 years, more than 90% showed reactivity to the anti-HAV, absence of infection markers for Hepatitis B and C, respectively HBsAg and anti-HCV, and a rate of 16.7% (95% CI 12.6-21.3) for the anti-HBs. The prevalence of neutralizing antibodies against the measles virus and yellow fever virus was 96.7% (95% CI 93.9-98.3) and 91.4% (95% CI 88.0-94.7) respectively. The polio results showed that 62.2% (95% CI 56.5-67.6) 71.7% (95% CI 66.2-76.6) and 63.5% (95% CI 56.5-69.6) had antibodies against the types 1, 2 and 3, respectively, showing vulnerability to B Hepatitis, Yellow Fever and Poliomyelitis.Item Epidemiologia molecular dos vírus dengue em Goiânia-GO, 1994 - 2006: vigilância laboratorial e caracterização dos sorotipos circulares(Universidade Federal de Goiás, 2008-08-06) FÉRES, Valéria Christina de Rezende; MARTELLI, Celina Maria Turchi; http://lattes.cnpq.br/5867052489026059Nowadays, dengue constitute the major public health problem, because is relevant cause of illness and death between thousands people that resident in the tropical and subtropical regions in world. The dengue virus is classified as four serotypes (DENV-1, 2, 3 and 4) according to antigenic differences and characterized intra-typical groups called genotypes. The laboratorial surveillance enables the diagnostic confirmation of dengue infection and monitoring serotypes circulating through the routine diagnostic techniques. Recently, the use molecular techniques has contributed to characterize and monitoring of the genotypes potentially virulent during epidemic and knowledge of biology of dengue virus. This thesis was organized in an introduction section, that include a literature review on dengue, and two manuscripts that describing the research conducted with focus on laboratory diagnostic and molecular epidemiology. The first manuscript entitled Laboratorial Surveillance of Dengue Virus in Central-Brazil, 1994-2003, was published at Journal of Clinical Virology, 2006 37 (3): 179-83. In this study we present the results of the virological surveillance for dengue cases conducted in the city of Goiânia (~1,200,000 population) from 1994 to 2003. Suspected cases were from the main public infectious disease reference hospital and outpatient clinics covering the metropolitan area. Serological and virus isolation tests were conduced at the regional reference laboratory. Our objective was to report dengue circulating serotypes from 1994 to 2003 and the role of distinct serotypes on dengue clinical outcomes in Central Brazil and to characterize serotypes and genotypes by reverse transcriptase PCR (RT-PCR) and by restricted site-specific PCR (RSS-PCR) patterns in selected samples. Laboratory surveillance identified mainly DEN-1 serotype from 1994 to 2002 shifting to a high circulation of DEN-3 in 2003. The adults (87,4%) were the most affected group and dengue fever accounted for the majority of the cases. Diagnosis of dengue was confirmed in ~50% of the suspected and enhanced by RT-PCR. RSS-PCR patterns for DEN-1 and DEN-3 corresponded to the circulating subtypes in the country. The infection DENV-3 did not suggest a major role of infecting DEN-3 in increasing disease severity during its first-year spread in Central Brazil. The second manuscript, to be submitted for publication is entitled: Epidemiologia Molecular do Vírus Dengue tipo 3 em Goiânia GO, 2005-2006. The objective of this manuscript was to characterize the DENV-3 genotype circulating isolated from well-characterized clinical and laboratory samples in Goiânia-GO/Brasil. Seven samples had sequences of the prM/M/E region obtained and comparative analysis was performed with the reference strains. The results showed the homology of the genomics sequences with genotype III strains. The nucleotide identity of the all the samples varied from 97.0% to 99.6% and the amino acid sequences from 97.5% to 99.5%. The analysis of the nucleotide sequence revealed silent mutation and 14 amino acid changes in the protein deduced from gene prM/M/E. In conclusion, the study confirms that the strains of DENV-3 in Central Brazil relate to genotype III. The genomic changes along Domain III of the protein E were observed, which could affect the pathogenicity, but were not consistent between samples of DCC and DHF. Samples of patients with dengue fever had mutations related to viral attenuation. More investigation is necessary to evidence of genomic changes found in relationship with clinical forms.Item Caracterização funcional da proteína Triose fosfato isomerase de Paracoccidioides brasiliensis como potencial adesina(Universidade Federal de Goiás, 2008-09-04) Pereira, Luiz Augusto; Soares, Célia Maria de Almeida; http://lattes.cnpq.br/8539946335852637; Ulhoa, Cirano José; Campos, Ivan Torres Nicolau; Izaac, Silvia Maria Salem; Giannini, Maria José Soares Mendes; Soares, Célia Maria de AlmeidaParacoccidioides brasiliensis, an important human pathogen causative of paracoccidioidomycosis (PCM), a systemic mycosis with broad distribution in Latin America. Adhesion to and invasion of host cells are essential steps involved in the infection and dissemination of pathogens. Furthermore, pathogens use their surface molecules to bind to host extracellular matrix components to establish infection. An adhesin of P. brasiliensiswas isolated from two dimensional electrophoresis and characterized. Peptides obtained by partial sequencing of the isolated protein, which presenteda molecular mass of 29 kDa and pI 5.8, were subjected to sequence analysis of their amino acids, that revealed strong homology to triose phosphate isomerase (TPI) from several sources. The complete cDNA and gene encoding TPI of P. brasiliensis (PbTPI) were characterized and both contained an open reading frame predicted to encode a 249 amino acid protein that presented all the peptides characterized in the native PbTPI. The complete coding PbTPI cDNA was cloned and over expressed in Escherichia coli host. The purified recombinant TPI was used to produce polyclonal antibody in rabbit. By immunoelectron microscopy and Western blot analysis, TPI was detected in the cell wall and the cytoplasm of the yeast phase of P. brasiliensis. The expression of PbTPI was analyzed in transition from mycelia to yeast phase. The native PbTPI is preferentially expressed in the yeast parasitic phase of P. brasiliensis. The recombinant PbTPI was found to bind to laminin and fibronectin in ligand far-Western blot assays. TPI binds preferentially to laminin, as determined by peptide inhibition assays. Of special note, the treatment of P. brasiliensisyeast cells with anti-PbTPI polyclonal antibody and the incubation of pneumocytes and VERO cells with the recombinant protein promoted inhibition of adherence and internalization of P. brasiliensisto those in vitrocultured cells. These observations indicate that TPI could be contribute to the adhesion of the microorganism to host tissues and to the dissemination of infection.Item Estudo dos protozoários intestinais oportunistas através da reação de polimerase em cadeia (PCR), em Goiânia-GO, Brasil (1999-2007)(Universidade Federal de Goiás, 2008-12-10) Souza Júnior, Edson Sidião de; García-Zapata, Marco Tulio Antonio; http://lattes.cnpq.br/3672512339058369; Garcia-Zapata, Marco Túlio Antonio; Gomes, Abraão Garcia; Anunciação, Carlos Eduardo; Lacerda, Elisângela de Paula Silveira; Carneiro, José RobertoThe Coccidia (Phylum Microspora) and Microsporidia (Phylum Apicomplexa) are responsible for a range of clinical pictures, and with important mortality rate, usually in patients with compromising of the immunological system. Its occurrence has been underestimated in our milieu maybe for the lack of knowledge on the part of the professionals of the health, or, by existence of a few qualified laboratories for its identification. The recognition of the species causing the infection is fundamental to guide the therapeutic handling to be adopted and definition of the infected patient's prognostic. This recognition is only possible through electronic Microscopy and of biomolecular techniques, standing out the of the Polymerase Chain Reaction (PCR). The present study it is part of a longitudinal study, developed in the IPTSP/UFG, on the clinical profile, epidemiologist and laboratorial of these agents. It specifically aims at to describe the standartization and the use of the PCR in the laboratorial diagnosis of opportunist protozoa in clinical samples human beings and as instrument of study of consumption waters human being in the city of Goiânia-GO, basic instrument for the control of these protozoa. The study it was carried through between october 1999 and december 2007. In the total were appraised 994 patient: 1) 664 immunocompromissed patients (with immunossuppression or immunodepression) with diarrhea coming from the School Hospital of UFG and of reference units in attendance to the health of the Goiás State; 2) 330 individuals from the community, by aleatory selection seemingly without disturbances of immunological systems and coming of the health reference units (unique sample). All of the samples were submitted to techniques of HPJ, Rugai and specific colorations for the diagnosis of the Coccidia (Hot Kinyoun) and intestinal Microsporidia (Hot-Chromotrope-Kokoskin), with previous coprological concentration through the ethyl formalin-acetate technique. The suggestive laboratorial infections were confirmed by the PCR technique, when the sample was enough. The confirmation and the identification of the species for PCR indicated the presence of the Encephalitozoon intestinalis, Enterocytozoon bieneusi, Cryptosporidium parvum/hominis in the analyzed samples. The standardization of the technique of PCR for ambient samples (water) was made from a survey carried through in the rivers and lakes of the city of Goiânia-GO. Inside of this quantitative one was found of Cryptosporidium parvum/hominis, demonstrating the viability of the PCR for ambient analyses that search the presence of opportunist protozoa in the water. These findings reveal, on one side, that these agents are present in ours milieu causing infections isolated or associated (co-infections), among them or with other infect-parasitic agents, being constituted in a risk for the populations of immunocompromised patients; and for other hand, they suggest in the need of implanting, more improvement diagnosis techniques, to define the clinical and epidemiological aspects of opportunistic protozoa species circulating in the Goiás State.Item FATORES DE RISCO E EPIDEMIOLOGIA MOLECULAR DE Streptococcus pneumoniae NÃO SUSCETÍVEIS À PENICILINA ISOLADOS DE NASOFARINGE DE CRIANÇAS QUE FREQUENTAM CRECHES EM GOIÂNIA-GO, BRASIL(Universidade Federal de Goiás, 2009-02-17) FRANCO, Cáritas Marquez; ANDRADE, Ana Lúcia Sampaio Sgambatti de; http://lattes.cnpq.br/7770363683068899Objectives: (i) to identify risk factors for S. pneumoniae penicillin nonsusceptible isolates (PNSp) in children attending day-care centers (DCCs) in Goiânia, Brazil and to assess the genetic patterns of pneumococcal isolates; (ii) to estimate the coverage for carriage serotypes for the 7-valente (PCV7) pneumococcal conjugate vaccine, and for the investigational 10 (PCV10) and 13-valent (PCV13) vaccines; (iii) to assess the genetic relatedeness between isolates expressing capsular type 14 and those non(sero)- typeable isolates (NTPn); (iv) to investigate if carriage isolates match genetically to any international pneumococcal clone (PMEN network). Methods: A cross-sectional survey of carriage PNSp was conducted among 1.192 children, 2 months to 5 years of age, attending 62 DCCs in Central Brazil. Capsular typing was performed in PNSp isolates (CLSI, 2007) and in a sample of isolates susceptible to penicillin (PSSp) matched to PNSp and DCCs whenever possible. Serotyping was performed by Quellung reactions and confirmed by multibead assay. NTPn isolates and serotype 14 were tested by PCR for capsule genes. Odds ratio for PNSp carriage and respective 95% confidence interval (95%CI) were assessed by logistic regression. Pulsed field gel electrophoresis (PFGE) was applied to assess the genetic similarity between PNSp serotype 14 and NTPn isolates. PCR was performed for the presence of pneumococcal capsule gene locus. For comparison purpose we also evaluated the genetic profile of PNSp serotype 14 invasive strains derived from the current pneumococcal invasive disease surveillance for the same pediatric population. Isolates were epidemiologically related if they shared ≥80% similarity on the dendrogram (Dice coefficient). A cluster was defined as three or more related isolates. Results: A total of 686 pneumococci were isolated for a colonization rate of 57.6% and 178 (25.8%) were PNSp. Among the PNSp isolates the usual common types were found: 14 (53%), 23F (10.2%), 6B (6%), 19F (4.8%) and 19A (4.2%). PSSp isolates displayed 30 different serotypes although serotype 14 was the most common. Overall a high prevalence of NTPn (11.1%) was observed with 62.9% PNSp. Serotypes coverage xvi for the PCV7, PCV10 and PCV13 vaccines were 55.2%, 55.9% and 65.1%, respectively. Being less than 24 months of age (OR=1.79; p=0.006), hospitalization in the previous three months (OR=2.19; p=0.025), and recurrent acute otitis media (OR=2.89; p=0.013) were independently associated with PNSp in a multivariate model. Among the 123 PNSp submitted to PFGE (106/carriage and 17/ invasive isolates) a major group of 34 serotype 14 strains (8 invasive and 26 carriage) was identified and found to be genetically related to the global pneumococcal clone Spain 9V-3 (82.7% similarity). All NTPn presented capsule gene locus and 10 (45.4%) of them presented capsule gene locus to type 14. Conclusions: (i) DCC attendees with history of recurrent AOM could significantly contribute to the spread of nasopharyngeal PNSp strains into the community; (ii) epidemiologic and molecular evidences support the findings that pneumococcal nonypeable carriage isolates are genetically similar to carriage and invasive isolates expressing capsular type 14; (iii) carriage and invasive isolates circulating in Goiânia belong to a serotype 14 variant of the Spain 9V -3 clone and play a critical role in the spread of PNSp strains to the entire pediatric community of GoiâniaItem Identificação e caracterização de moléculas envolvidas na interação de Paracoccidioides brasiliensis com o hospedeiro(Universidade Federal de Goiás, 2009-03-31) DANTAS, Sabrina Fonseca Ingênito Moreira; SOARES, Célia Maria de Almeida; http://lattes.cnpq.br/8539946335852637Paracoccidioides brasiliensis causes paracoccidioidomycosis (PCM), a systemic mycosis presenting clinical manifestations ranging from mild to severe forms. A P. brasiliensis cDNA expression library was produced and screened with pooled sera from PCM patients adsorbed against antigens derived from in vitro-grown P. brasiliensis yeast cells. Sequencing DNA inserts from clones reactive with PCM patients sera indicated 35 open reading frames presenting homology to genes involved in metabolic pathways, transport, among other predicted functions. The complete cDNAs encoding aromatic L-amino acid decarboxylase (Pbddc), lumazine synthase (Pbls) and a homologue of the high affinity copper transporter (Pbctr3) were obtained. Recombinant proteins PbDDC and PbLS were obtained; a peptide was synthesized for PbCTR3. The proteins and the synthetic peptide were recognized by sera of patients with confirmed PCM and not by sera of healthy patients. Using the vivo-induced antigen technology (IVIAT) we identified immunogenic proteins expressed at high levels during infection. Quantitative real - time RT-PCR demonstrated high transcript levels of Pbddc, Pbls and Pbctr3 in yeast cells infecting macrophages. Transcripts in yeast cells derived from spleen and liver of infected mice were also measured by qRT-PCR. Our results suggest a putative role for the immunogenic proteins in the infectious process of P. brasiliensis.Item Investigação epidemiológica e molecular da infecção pelo vírus da hepatite C em usuários de drogas ilícitas no Brasil Central(Universidade Federal de Goiás, 2009-04-02) LOPES, Carmen Luci Rodrigues; MARTINS, Regina Maria Bringel; http://lattes.cnpq.br/2582896795892370Hepatitis C virus (HCV) infection is an important problem of public health. Drug users (DU) constitute a group of frequent exposure to HCV and little is known about this infection in DU in Brazil. This study aimed to investigate the seroepidemiological and molecular profile of HCV infection among drug users in Central Brazil. A total of 691 DU, being 102 injection drug users (IDU) and 589 noninjecting drug users (NIDU), were interviewed and blood samples collected in 26 treatment drug centers in Campo Grande-MS and Goiânia-GO. Blood samples (sera) were tested for antibodies to HCV (anti-HCV). Anti-HCV-positive samples were submitted to HCV RNA detection by polymerase chain reaction (PCR) with primers complementary to the 5 NC and NS5B regions of viral genome and genotyped by LiPA and direct nucleotide sequencing followed by phylogenetic analysis, respectively. The anti-HCV prevalence was 6.9% (95% CI: 5.2-9.2). The study population reported a low known regarding HCV transmission ways, such as parenteral (20.8-30.5%), sexual (31,7%) and vertical (20%) of HCV. Multivariate analysis of risk factors revealed that age > 40 years, route (injecting) and duration of drug use and blood transfusion were associated with HCV infection. HCV RNA was detected in 85.4% of the anti-HCV-positive samples. Thirty-three samples were of genotype 1 by the LiPA, subtypes 1a (63.4%) and 1b (17.1%), and eight samples (19.5%) were of genotype 3, subtype 3a. The phylogenetic analysis of the NS5B region showed that 17 (68%), 5 (20%) and 3 (12%) samples were of subtypes 1a, 3a and 1b, respectively. This study shows a high HCV infection prevalence and the predominance of subtype 1a among drug users in Central Brazil. Also, the injecting drug use was the risk factor most strongly associated with this infection. In addition, the study population reported a low known regarding HCV transmission ways. Thus, preventive measures are needed to control this infection in illicit drug usersItem Epidemiologia molecular e riscos associados ao portador nasal de Staphylococcus aureus isolados de crianças de creches de Goiânia(Universidade Federal de Goiás, 2009-04-15) CARDOSO, Juliana Lamaro; KIPNIS, André; http://lattes.cnpq.br/4434965360286741; ANDRADE, Ana Lúcia Sampaio Sgambatti de; http://lattes.cnpq.br/7770363683068899Objectives: (i) to assess the prevalence of Staphylococcus aureus and methicillin-resistant S. aureus (MRSA) nasal carriage in children attending day-care centers (DCC) in the municipality of Goiânia; (ii) to determine the potential risk factors related to S. aureus carriage and MRSA; (iii) to characterize MRSA isolates circulating in DCCs using molecular typing methods. Methods: Between August and December 2005, nasal swabs were collected from children who attended 62 DCCs. Clinical and socio-demographic information associated with the acquisition of S. aureus and MRSA were obtained through questionnaires applied to parents or guardians. The swabs were processed following the standard methods for identification and isolation of S. aureus. Amplification femB gene by polymerase chain reaction (PCR) was used to confirm the specie. The presence of mecA gene was detected by PCR and the positive isolates were identified as MRSA. Susceptibility to MRSA was determined by disk diffusion method. MRSA molecular typing was performed by PFGE, MLST, spa typing and SCCmec multiplex PCR. Results: 371 (31.1%) out of the 1.192 collected swabs were positive for S. aureus and 14 (1.2%) were identified as MRSA. The factors independently associated with risks for nasal colonization by S. aureus were children higher than two years of age (OR = 1.83, 95% CI 1.27-2.65) and previous DCC attendance (OR = 1.48; 95% CI 1.01-2.16). Mother s high degree of education was a protective factor for S. aureus carriage (OR = 0.43, 95% CI 0.23-0.80). A multidrug resistant dominant MRSA lineage was identified comprising 8 out of the 14 MRSA isolates. This cluster was characterized as SCCmec type IIIA, ST239 and spa type t037 sharing 82.7% genetic similarity with the Brazilian clone. One MRSA strain was classified as SCCmec type V and ST1120. This strain showed features of CA-MRSA although it has been recovered from a healthy child who presented risk factors for HA-MRSA acquisition. The remaining MRSA strains showed a diverse genetic background. Conclusions: Children attending DCCs are often colonized with S. aureus and although the prevalence of MRSA was low, they can represent potential vectors of spread of resistant pathogens to the community. The detection of a MRSA lineage circulating within DCCs suggests a two-way flow spread of MRSA between hospitals and community.Item Detecção de vírus gastroentéricos em mulheres em Goiânia-GO(Universidade Federal de Goiás, 2009-04-30) FERREIRA, Rui Gilberto; CARDOSO, Divina das Dôres de Paula; http://lattes.cnpq.br/9770835116155857The gastroenteric viruses are important etiological agents of gastroenteritis in individuals of all ages. It is believed that individuals with deficits in the immune system (humoral and/or cellular), among them pregnant women and human immune deficiency virus (HIV)-seropositive women, are more susceptible to these viral infections. The rotaviruses, adenoviruses, astroviruses, and caliciviruses constitute are among the main causes of acute gastroenteritis in the world, and are accounted for high morbi-mortality rates, especially among children under five years of age. It is believed that, by the age of three, approximately 90% of all children in developing countries have antibodies to one or more of these agents. Neonatal infection does not exclude the possibility of re-infection, with different viral serotypes, however it protects the individual against severe disease. This study aimed at the detection of rotavirus, adenovirus, astrovirus e calicivirus in women seeing at the Hospital das Clínicas da Universidade Federal de Goiás (HC-UFG) and at the investigation of as association between the positivity to this viruses and the low immune status, characteristic in pregnant women and/or HIV-seropositive women. This was a prospective follow-up study of women seeing at the Gynecology and obstetrics (OB-GYN) sector of the HC-UFG aiming at the detection of gastroenteric viruses (rotavirus, adenovirus, astrovirus e calicivirus). For this, fecal samples were collected from 84 women, in the period from July-2006 to June-2007. For rotavirus detection, fecal samples were screened by polyacrilamide gel electrophoresis (PAGE) and by an immunoenzimatic assay (IEA). The calicivirus and astrovirus were detected by polymerase chain reaction post-reverse transcription (RT-PCR), and adenovirus detection was performed by an EIE. The astrovirus genotyping was conducted by Nested-PCR. Three-hundred and fourteen fecal samples were collected from a total of 84 women. From those 84 women, 29 were HIV-seropositive, 55 HIVseronegative, 45 were pregnant at the time, and 39 were not pregnant. The patients were aged between 16 and 67 years-old, and 47% of them had up to 30 years of age. From the total 84 patients, 19 (22.6%) were positive for calicivirus and/or astrovirus in at least one of the collected samples, as follows: calicivirus (14/19) and astrovírus (6/19), with the highest positivity rates being detected in the months of July and August (astrovirus) and September and October (calicivirus). None of the collected samples were positive for rotavirus or adenovirus. The association index between gastroenteric virus positivity and pregnancy, in the presence or not of HIV-seropositivity, was of 68.4% (13/19); however, there was no significant difference between the group of women that were not pregnant and the ones that were HIV-seronegative. The gastroenteric viruses were detected in a significant parcel (22.6%) of this population of adult women, and were represented by the caliciviruses (16.7%) and astroviruses (7.1%). Under the conditions of the present study, no association was found between pregnancy and/or positivity for HIV, as reducing factors for the immunological capacity of the women, and the detection of gastroenteric viruses; in conclusion, the pregnancy and/or HIV-seropositivity did not increase the chances of these women to be infected by these gastroenteric viruses.Item Marcadores moleculares, imunológicos e genéticos das reações hansênicas(Universidade Federal de Goiás, 2009-05-08) SOUSA, Ana Lúcia Osório Maroclo de; MARTELLI, Celina Maria Turchi; http://lattes.cnpq.br/5867052489026059; STEFANI, Mariane Martins de Araújo; http://lattes.cnpq.br/5581414958714905Type 1 (T1R) and Type 2 (T2R) leprosy reactions are complications in the clinical management of leprosy patients because they can lead to neural damage and impairment, resulting in irreversible deformities and disabilities. This thesis, presented as research article/manuscript has investigated potential markers for the diagnosis and prognosis of leprosy reactions. In the first study, we evaluated a multicentric cohort of leprosy patients with single skin lesion which is considered the earliest clinical manifestation of the disease. In this study, at the moment of diagnosis a skin biopsy was collected for histopathology and for the investigation of Mycobacterium leprae DNA by polymerase chain reaction (ML-PCR). After diagnosis patients were treated with single dose of Rifampicin, Ofloxacin and Minocyclin (ROM) and were clinically monitored during 3 years .During follow up, around 15% of patients developed T1R. In multivariate analysis, age > 40 years and MLPCR positivity were associated with T1 manifestation. The second study aimed to identify potential circulating markers associated with leprosy reactions. Plasma levels of 27 cytokines/chemokines/growth factors were quantified by multiplex assay. A nested case control study compared the levels of these factors in leprosy patients with or without T1R and T2R. Leprosy patients were paired by sex, age group (+/- 5 years) and histopathological classification. Significant differences in plasma levels of CXCL10 (p=0.004) and IL6 (p=0.013) were observed in patients with T1R compared with controls without reaction. IL7 levels (p=0.039) and PDGF-BB (p=0.041) were increased in T2R and marginally significant for IL6 (p=0.05). This investigation indicated that CXCL10, IL6 and IL7 may represent potential plasma biomarkers of leprosy reactions. The third study investigated the influence of single nucleotide polymorphism (SNP) in the IL6 gene and the development of leprosy reactions. For this purpose a nested case control study was performed based on a cohort of 409 leprosy patients recruited in Goiânia-GO. After leprosy diagnosis patients were monitored during multidrug therapy regarding the appearance of leprosy T1R and T2R. Evidences of positive associations were observed for leprosy T2R and tag SNPs markers- rs 2069832 (p=0.002), rs 2069840 (p=0.027) and rs 2069845 (p=0.044). These IL6 gene tag SNPs capture information of the whole gene locus. Positive association with T2R was also seen for the functional variant of IL6 gene- tag SNP rs 1800795 (p=0.005). Additionally, IL6 plasma levels among TR2 patients and controls without reaction correlated with different IL6 genotypes. No association was observed between IL6 gene variants and leprosy T1R. The description of genetic predictive factors of leprosy reactions may contribute to the development of preventive strategies against these leprosy incapacitating events.Item Estudo das alterações morohistológicas em larvas de Aedes aegypti submetidas ao tanino catéquino isolado da Magonia pubescene (Sapindaceae) e ao diterpeno de Copaifera reticulata(Leguminosae)(Universidade Federal de Goiás, 2009-06-25) VALOTTO, Cleyde Ferreira Barreto; SILVA, Ionizete Garcia da; http://lattes.cnpq.br/5021551669347602The dengue and yellow fever are caused by Flavivirus, and are an importants problems in public health in the range of tropical and subtropical world. There is only available vaccine for yellow fever. The transmission of both diseases to humans is by the bite of female Aedes aegypti. The vector control is done through the elimination of breeding potential, application of larvicides in collections of water for adults vectors space applications of insecticides. The problems arising from continuous use of chemical insecticides have been the emergence of resistant populations to these products and concern about the environmental damage caused by slow degradation. In search of alternatives for the vector control, studies are being developed with substances of botanical origin, with the aim to control the vectors and preserve the environment. The use of natural insecticides has some advantages when compared to synthetic because they are renewable and have rapid degradation. Moreover, its composition is quite complex, reducing the speed of emergence of resistance and be of low toxicity to mammals, birds and environment. This study presents the morphohistologic and ultrastructural changes of catechist tannin extracted from Magonia pubescens, and the Diterpene extracted from Copaifera reticulata on the larvae of A. aegypti with the objective to evidence the mechanisms of action of these substances. For light microscopy, the 3rd stage larvae were subjected to a solution of catechist tannin 37ppm and the diterpene 9ppm, where they remained for up to 24h. Larvae that reached lethargic state were collected and fixed in 4% in the paraformoldehyde buffer 0.1 M sodium cacodilate pH 7.2, included in resin and the slides stained with hematoxylineosin technique. For electron microscopy, larvae were subjected to the same concentrations of catechist tannin and diterpene, where they remained for 22 h. Larvae that reached lethargic state were collected and dissected and their gut fixed in 2% glutaraldehyde, 2% paraformoldehyde, 3% sucrose in sodium buffer cacodilate 0.1 M pH 7.2 and post fixed in osmium tetroxide at 1% in cacodilate sodium 0.1 M. After this they were contrasted with uranyl acetate 1%, dehydrated, embedded and polymerized. The ultra sections were contrasted with uranyl acetate 3% and lead citrate and were caracterized by eletron microscopy. The changes caused by catechist tannin observed by light 20 and electron microscopy were high vacuolation, absence of cytoplasmic limits, cellular aging, cell disruption, vesicular apical formation with release of cytoplasmic content, increase in intercellular spaces, detachment of cells from the basal lamina and structural change of microvilli. Changes by diterpene by light and electron microscopy were high cytoplasmic vacuolation, cells and nuclei hypertrophy, degeneration of the edge brush, vesicular apical formation with release of cytoplasmic contents of cells, stratification of the epithelium and folds in the peritrophic matrix.Item Surto de infecção após videoscopias causado por Mycobacterium massiliense em Goiânia-GO : análise molecular e determinação da suscetibilidade aos antimicrobianos(Universidade Federal de Goiás, 2009-12-03) CARDOSO, Alessandra Marques; KIPNIS, Ana Paula Junqueira; http://lattes.cnpq.br/1252262903952987; KIPNIS, André; http://lattes.cnpq.br/4434965360286741In recent years the number of infections caused by microbacteria non-tuberculous mycobacteria (NTM) has increased mainly due to opportunistic infections in individuals imunocompormetidos and improvement of farming techniques and identification of MTN. Mycobacterium massilienese is an emerging body associated with wound infections, abscesses and pneumonia. An outbreak of infection after videoscopy occurred between 2005 and 2007 in seven hospitals in Goiânia-GO, in central Brazil. The objective of this study was to identify NTM isolated from patients with infection after arthroscopy and lararoscopia by PCR followed by analysis of fragment length polymorphism restrção (PRA-hsp65), compared by gel electrophoresis pulsed-field gel (PFGE), sequencing of the partial rpoB gene and determination of antimicrobial susceptibility in vitro. NTM were recovered from samples (exudate abscess subcutâneio) of 18 patients involved in the outbreak. In the period leading up to this study there was no reported case of infection after videoscopy caused by MTN in Goiania. The 18 isolates were identified as M, massiliene and genotyped as a single clone, indicating that they had a common origin, suggesting a common source of infection for the patients involved in the outbreak. The epidemic isolates were susceptible to amikacin (MIC90 4 micrograms / ml) and clarithromycin (MIC90 <1 ug / ml), but resistance to ciprofloxacin (MIC90 <128g/ml), tobramycin (MIC90 32 micrograms / ml) and intermediate susceptibility to cefoxitin (MIC90 64 ug / ml). In conclusion this study demonstrated the clonality of strains of M. massiliense involved in infections after procedures videoscopes and that they are susceptible to drugs indicated for the treatmentItem Tipagem molecular de Streptococcus pneumoniae isolados da nasofaringe de crianças no contexto da vacinação pneumocócica(Universidade Federal de Goiás, 2010-02-18) ROCHA, Cristyane Gonçalves Benicio Bastos; ANDRADE, Ana Lúcia Sampaio Sgambatti de; http://lattes.cnpq.br/7770363683068899; PIMENTA, Fabiana Cristina; http://lattes.cnpq.br/2230554075502158Objectives (i) Present review article focusing on pneumococcal vaccines and carriage; (ii) to validate sequential multiplex PCR for identifying pneumococcal capsular serotypes from children attending day-care centers; (iii) determine the multilocus sequence typing; (iv) to identify the capsular types of multiple colonies of S. pneumoniae isolates from a single sample of nasopharyngeal secretions of children attending day-care centers in Goiânia. Materials and Methods S. pneumoniae was obtained from health children less than 5 years old attending 62 day care centers of Goiânia. The laboratory procedures were performed according to WHO recommendations. Were selected 217 isolates (penicillin resistant and sensitive) for capsular typing by multiplex PCR technique. MLST was performed for 55 isolates representing the serotypes detected and the different susceptibility patterns for penicillin. Quellung reaction was used for typing isolates serotypes 6A, 6B, 18C and the isolates not typed by multiplex PCR. For 28 presumptive pneumococcal positive NP swabs, 3 colonies were picked to acess possible serotype diversity. Eighty four pneumococci were identified by conventionally procedure and multiplex PCR was performed. Results Serotypes were deduced for 177/217 (81.6%) of the pneumococcal. The most frequent serogroups/serotypes were 14, 6, 23F, 19F and 18. Multiple serotypes were detected in 13 specimens. Were found 19 MLST types and two new ST. Forty (18.4%) were not serotyped by the multiplex PCR and quellung reaction. The analysis of three colonies from the same NP permitted the detection of differente serotypes in 7/28 (25%) NP samples. Conclusion (i) The multiplex PCR is simple and cost-effective method for detecting multiple serotypes in nasopharyngeal isolates; (ii) and thus might be useful for the monitoring of pneumococcal colonization over time; (iii) the use of multiplex PCR can further broaden our understanding of the dynamics of pneumococcal carriage, including multiple serotypes, the effect of vaccination on carriage, and transmission, as well as surveillance of IPD and co-colonization.Item Caracterização morfológica, molecular e biológica de fungos patogênicos a invertebrados dos Cerrados de Goiás(Universidade Federal de Goiás, 2010-02-23) ROCHA, Luiz Fernando Nunes; KIPNIS, André; http://lattes.cnpq.br/4434965360286741; LUZ, Wolf Christian; http://lattes.cnpq.br/1104009511235835The high biodiversity of fungi pathogenic to invertebrates and their potential to control pests until today not well known emphasize the importance to look for new effective species and strains. The Cerrado is considered one of the hotspots of biodiversity and little is known about the occurrence and the potential of fungi pathogenic to invertebrates found in this biome. In the present study a survey of fungi was carried out in different areas of the Cerrado of Goiás. Samples of soil, slurry, water and moribund or dead insects were collected for isolation of fungi. Vectors of medical importance such as triatomines (Triatoma infestans and Rhodnius neglectus), mosquitoes (Aedes aegypti and Culex quinquefasciatus), ticks (Riphicephalus (Boophilus) microplus) and snails (Biomphalaria glabrata) were used as baits for isolation of fungi. After isolation fungi were morphologically identified and included in the collection ofInstitute of Tropical Pathology and Public Health, Federal University of Goiás. Some isolates of Evlachovaea and Metarhizium were molecularly characterized and activity tested against T. infestans. A total of 561 samples of soil (440), slurry (106) and water (15) was collected in different areas of Goiás State. Concerning samples collected at Fazenda Santa Branca, 68 isolates of pathogenic fungi were obtained and identified as belonging to the genera Aspergillus, Beauveria, Cladosporium, Evlachovaea, Fusarium, Gliocladium, Isaria,Lecanicillium, Metarhizium, Paecilomyces, Pochonia and Trichoderma. An total of 106 isolates of Metarhizium spp. and 6 of Evlachovaea spp. were sampled in other areas, being 65isolates of Metarhizium and 1 of Evlachovaea from the Ema National Park, 33 and 1 from the Northern portion of Goiás state, and 8 and 4 from the Silvânia National Forest, respectively.Most fungi were isolated from soils using triatomines as baits. Fungi from genera Aschersonia, Batkoa, Beauveria, Cordyceps, Evlachovaea, Fusarium, Lecanicillium, Pandora and Torrubiella were isolated from mycosed cadavers. All tested isolates of Metarhizium spp. and Evlachovaea spp. induced high mortality of T. infestans in relative humidity (RH) close to saturation. The lowest values for lethal time of 90% were 6.6 d (6.3 7.1 d; M. robertsii IP 34)and 7.1 d (6.7 7.8 d; Evlachovaea IP 141), after treatment of T. infestans and exposure to RH> 98%. The lethal concentration to obtain 50% mortality (LC50) of IP 34 was 2.8x103 (C.I. 4.4x102-4.6x103) and the LC90 was 7.2x103 (C.I. 4.4x103-6.4x105) CFU/cm2 at 10 d p.i. In RH 75% mortality of triatomines did not exceed 20%. Morphological studies and sequencing of the ITS and TEF region of Evlachovaea isolates showed that genus Evlachovaea must be synonymized with Isaria, and that the largest group of isolates previously identified as Evlachovaea are I. cateniannulata, whereas the smaller group is probably a new species of Isaria. The sequencing of the TEF and ITS regions showed that Metarhizium isolates belong to species M. anisopliae, M. robertsii, M. flavoviride var. pemphigi, and the largest group of Metarhizium isolates can be a new species of Metarhizium or a M. anisopliae variety. The results confirmed that in the Cerrado a high diversity of fungi is present and some of them, in special M. robertsii (IP 34) and Evlachovaea (IP 141) have potential for biological control of T. infestans