Análise dos polimorfismos genéticos dos genes msp1 e csp em isolados de P. vivax em Porto Velho – Rondônia

Dissertação apresentada ao Programa de Pós- Graduação: Mestrado em Biologia Experimental (PGBIOEXP) da Fundação Universidade Federal de Rondônia (UNIR) como requisito final para a obtenção do título de Mestre em Biologia Experimental. Orientador: Prof. Dr. Mauro Shugiro Tada.A malária vivax apresenta uma aparente evolução benigna. Embora esta evolução seja branda, sua morbidade é elevadíssima nas regiões endêmicas, de modo que, fora da África, a incidência de P. vivax é superior a de P. falciparum. Vários motivos podem explicar estes índices elevados, como a recaída, fluxo migratório, falha terapêutica e resistência de cepas. A menos que um plano de combate à doença considere estes fatores, seus objetivos malograrão, como em várias propostas de erradicação. Assim, novas estratégias foram baseadas em particularidades locais, de modo que, no Brasil, houve uma redução da prevalência da malária, contudo sua incidência continua alta. Por conseguinte, novas estratégias de combate à malária, principalmente a vivax, devem levar em conta conhecimentos mais específicos à cada localidade, como por exemplo, a dinâmica de sua transmissão. Com este intuito, vários estudos sobre polimorfismos gênicos foram realizados, para distinguir populações de parasitos, especialmente utilizando os genes da proteína de superfície do merozoíto (msp1) proteína circunsporozoíta (csp). Utilizamos esta estratégia para analisar a diversidade de cepas de P. vivax no município de Porto Velho oriundas de amostras de áreas urbanas e rurais. Para tal, foi analisada 224 amostras, aproximadamente 70% de áreas rurais, a partir das quais foram extraído DNA plasmodial. Utilizamos iniciadores de regiões polimórficas I, II e III no gene msp1 e um bloco polimórfico do gene csp, as regiões de interesse foram amplificadas por PCR. Foram detectados, aproximadamente, 158 isolados, em média. A diversidade genética observada foi similar na área rural (He = 0,7919) e urbana (He = 0,8112) em média, contudo, não foi verificado diferenças estatisticamente significantes entre as amostras. Nas regiões I, II e III do gene msp1 foram detectadas 50, 10 e 12 infecções policlonais, respectivamente, ao passo que no sistema CPS foi detectado 43. Os resultados indicam que os parasitos analisados de regiões rurais são, estatisticamente, semelhantes àqueles de áreas urbanas. Portanto, estes resultados podem indicar a possibilidade de circulação de parasitos entre áreas urbanas e rurais, de tal sorte que aumenta a possibilidade de que cepas resistentes, ou com maior virulência, possam espalhar rapidamente dentro do município

Epidemiological profile of Zika, Dengue and Chikungunya virus infections identified by medical and molecular evaluations in Rondonia, Brazil

Several arboviruses have emerged and/or re-emerged in North, Central and SouthAmerican countries. Viruses from some regions of Africa and Asia, such as the Zika and Chikungunya virus have been introduced in new continents causing major public health problems. The aim of this study was to investigate the presence of RNA from Zika, Dengue and Chikungunya viruses in symptomatic patients from Rondonia, where the epidemiological profile is still little known, by one-step real-time RT-PCR. The main clinical signs and symtoms were fever (51.2%), headache (78%), chills (6.1%), pruritus (12.2%), exanthema (20.1%), arthralgia (35.3%), myalgia (26.8%) and retro-orbital pain (19.5%). Serum from 164 symptomatic patients were collected and tested for RNA of Zika, Dengue types 1 to 4 and Chikungunya viruses, in addition to antibodies against Dengue NS1 antigen. Direct microscopy for Malaria was also performed. Only ZIKV RNA was detected in 4.3% of the patients, and in the remaining 95.7% of the patients RNA for Zika, Dengue and Chikungunya viruses were not detected. This finding is intriguing as the region has been endemic for Dengue for a long time and more recently for Chikungunya virus as well. The results indicated that medical and molecular parameters obtained were suitable to describe the first report of symptomatic Zika infections in this region. Furthermore, the low rate of detection, compared to clinical signs and symptoms as the solely diagnosis criteria, suggests that molecular assays for detection of viruses or other pathogens that cause similar symptoms should be used and the corresponding diseases could be included in the compulsory notification list

CHARACTERIZATION OF ENTEROAGGREGATIVE ESCHERICHIA COLI AMONG DIARRHEAL CHILDRENIN WESTERN BRAZILIAN AMAZON

ABSTRACT BACKGROUND: Enteroaggregative Escherichia coli (EAEC) is one of the main acute and chronic diarrhea causes both in children and adults, mainly in developing countries. OBJECTIVE: The aim of the present study is to characterize EAEC strains isolated from faecal samples and to identify genes potentially contributing to virulence, biofilm production and antimicrobial resistance in children admitted to a pediatric hospital in Porto Velho, Rondônia State. METHODS: The total of 1,625 E. coli specimens were isolated from 591 children in the age group 6 years or younger who were hospitalized in Cosme and Damião Children Hospital in Porto Velho, between February 2010 and February 2012, with acute gastroenteritis. Colonies suggestive of E. coli were subjected to polymerase chain reaction testing in order to identify the virulence factors. The in vitro adhesion assays using HEp-2 adherence were tests. Biofilm detection through spectrophotometry and antimicrobial susceptibility tests were conducted in the disk diffusion method. RESULTS: The mentioned study examined 591 stool samples from children with diarrhea. Diarrheogenic E. coli was found in 27.4% (162/591) of the children. EAEC was the diarreagenic E. coli most frequently associated with diarrhea 52.4% (85/162), which was followed by enteropathogenic E. coli 43.8% (71/162), enterotoxigenic E. coli 2.4% (4/162), and enterohemorrhagic E. coli 1.2% (2/162). The aggR gene was detected in 63.5% (54/85) of EAEC isolates; moreover, statistically significant correlation was observed among typical EAEC (aggR) and aatA (P<0.0001), irp2 (P=0.0357) and shf (P=0.0328). It was recorded that 69% (59/85) of the 85 analyzed EAEC strains were biofilm producers; 73% (43/59) of the biofilm producers carried the aggR gene versus 42.3% (11/26) of non-producers (P=0.0135). In addition, there was association between the aatA gene and biofilm production; 61% (36/59) of the samples presented producer strains, versus 19.2% (5/26) of non-producers (P<0.0004). Antibiotic sensitivity test evidenced that most EAEC were ampicillin 70.6% (60/85), sulfamethoxazole 60% (51/85), tetracycline 44.7% (38/85) and cefotaxime 22.4% (19/85) resistant. CONCLUSION: As far as it is known, the present study is pioneer in Northern Brazil to investigate EAEC virulence factors and to show the antimicrobial susceptibility of EAEC strains isolated from children with diarrhea

Chronic Cystoisospora belli infection in an HIV/AIDS patient treated at the specialized assistance service in Porto Velho County - Rondônia.

Abstract Cystoisospora belli infection manifests as diarrhea, and can potentially progress to malabsorption in HIV patients. Here, we report a case of C. belli infection in an HIV/AIDS patient with chronic diarrhea symptoms for at least 2 years. Coproscopic analyses based on direct technique and modified Ziehl-Neelsen technique without a commercial kit were performed. The current case report highlights the protocol to be adopted in coproscopic analyses applied to HIV patients. The importance of including the appropriate parasitological testing of patients with chronic intestinal isosporiasis in parasitological test routines must be considered

Cross-reactive anti-PfCLAG9 antibodies in the sera of asymptomatic parasite carriers of Plasmodium vivax

Submitted by EMERSON LEAL ([email protected]) on 2016-06-22T14:24:43Z No. of bitstreams: 1 Cross-reactive anti-PfCLAG9.pdf: 858856 bytes, checksum: 2ffe44623a0b886c451e6b51473c40a8 (MD5)Approved for entry into archive by EMERSON LEAL ([email protected]) on 2016-07-14T15:32:27Z (GMT) No. of bitstreams: 1 Cross-reactive anti-PfCLAG9.pdf: 858856 bytes, checksum: 2ffe44623a0b886c451e6b51473c40a8 (MD5)Made available in DSpace on 2016-07-14T15:32:27Z (GMT). No. of bitstreams: 1 Cross-reactive anti-PfCLAG9.pdf: 858856 bytes, checksum: 2ffe44623a0b886c451e6b51473c40a8 (MD5) Previous issue date: 2013Made available in DSpace on 2016-07-15T18:40:36Z (GMT). No. of bitstreams: 3 Cross-reactive anti-PfCLAG9.pdf.txt: 39798 bytes, checksum: 0239dd87d08632acfc502eb84feeeb5c (MD5) Cross-reactive anti-PfCLAG9.pdf: 858856 bytes, checksum: 2ffe44623a0b886c451e6b51473c40a8 (MD5) license.txt: 2991 bytes, checksum: 5a560609d32a3863062d77ff32785d58 (MD5) Previous issue date: 2013Centro de Pesquisa em Medicina Tropical. Porto Velho, RO, Brasil / Universidade Federal de Rondônia. Porto Velho, RO, Brasil.Fundação Oswaldo Cruz. Fiocruz Rondônia. Instituto de Pesquisas em Patologias Tropicais. Porto Velho, RO, Brasil.Centro de Pesquisa em Medicina Tropical. Porto Velho, RO, Brasil.Centro de Pesquisa em Medicina Tropical. Porto Velho, RO, Brasil.Fundação Oswaldo Cruz. Fiocruz Rondônia. Instituto de Pesquisas em Patologias Tropicais. Porto Velho, RO, Brasil.Centro de Pesquisa em Medicina Tropical. Porto Velho, RO, Brasil / Universidade Federal de Rondônia. Porto Velho, RO, Brasil.Centro de Pesquisa em Medicina Tropical. Porto Velho, RO, Brasil.Centro de Pesquisa em Medicina Tropical. Porto Velho, RO, Brasil / Fundação Oswaldo Cruz. Fiocruz Rondônia. Instituto de Pesquisas em Patologias Tropicais. Porto Velho, RO, Brasil.Fundação Oswaldo Cruz. Fiocruz Rondônia. Instituto de Pesquisas em Patologias Tropicais. Porto Velho, RO, Brasil.Centro de Pesquisa em Medicina Tropical. Porto Velho, RO, Brasil / Fundação Oswaldo Cruz. Fiocruz Rondônia. Instituto de Pesquisas em Patologias Tropicais. Porto Velho, RO, Brasil.The PfCLAG9 has been extensively studied because their immunogenicity. Thereby, the gene product is important for therapeutics interventions and a potential vaccine candidate. Antibodies against synthetic peptides corresponding to selected sequences of the Plasmodium falciparum antigen PfCLAG9 were found in sera of falciparum malaria patients from Rondônia, in the Brazilian Amazon. Much higher antibody titres were found in semi-immune and immune asymptomatic parasite carriers than in subjects suffering clinical infections, corroborating original findings in Papua Guinea. However, sera of Plasmodium vivax patients from the same Amazon area, in particular from asymptomatic vivax parasite carriers, reacted strongly with the same peptides. Bioinformatic analyses revealed regions of similarity between P. falciparum Pfclag9 and the P. vivax ortholog Pvclag7. Indirect fluorescent microscopy analysis showed that antibodies against PfCLAG9 peptides elicited in BALB/c mice react with human red blood cells (RBCs) infected with both P. falciparum and P. vivax parasites. The patterns of reactivity on the surface of the parasitised RBCs are very similar. The present observations support previous findings that PfClag9 may be a target of protective immune responses and raises the possibility that the cross reactive antibodies to PvCLAG7 in mixed infections play a role in regulate the fate of Plasmodium mixed infections