Infection of the central nervous system with dengue virus 3 genotype I causing neurological manifestations in Brazil

Abstract: A case of dengue virus 3 (DENV-3) genotype I infection with neurological manifestations occurred in Belo Horizonte, Minas Gerais in October 2012. The serotype was detected by PCR, and the genotype was assessed by sequencing and phylogenetic analysis of the C-prM region. The virus causing neurological manifestations clustered with other sequences of DENV-3 genotype I. Because neurological manifestations of DENV are possibly misdiagnosed in Brazil, this study serves as an alert of the importance of DENV diagnoses in CNS infections.

Identification of Leptospira serovars by RFLP of the RNA polymerase beta subunit gene (rpoB)

Leptospires are usually classified by methods based on DNA-DNA hybridization and the conventional cross-agglutination absorption test, which uses polyclonal antibodies against lipopolysaccharides. In this study, the amplification of the rpoB gene, which encodes the beta-subunit of RNA polymerase, was used as an alternative tool to identify Leptospira. DNA extracts from sixty-eight serovars were obtained, and the hypervariable region located between 1990 and 2500-bp in the rpoB gene was amplified by polymerase chain reaction (PCR). The 600-bp amplicons of the rpoB gene were digested with the restriction endonucleases TaqI, Tru1I, Sau3AI and MslI, and the restriction fragments were separated by 6% polyacrylamide gel electrophoresis. Thirty-five fragment patters were obtained from the combined data of restriction fragment length polymorphism (PCR-RFLP) analysis and used to infer the phylogenetic relationships among the Leptospira species and serovars. The species assignments obtained were in full agreement with the established taxonomic classifications. Twenty-two serovars were effectively identified based on differences in their molecular profiles. However, the other 46 serovars remained clustered in groups that included more than one serovar of different species. This study demonstrates the value of RFLP analysis of PCR-amplified rpoB as an initial method for identifying Leptospira species and serovars.

Mass trapping with MosquiTRAPs does not reduce Aedes aegypti abundance

The objective of this study was to evaluate the effectiveness of Aedes aegypti mass trapping using the sticky trap MosquiTRAP (MQT) by performing a cluster randomised controlled trial in Manaus, state of Amazonas, Brazil. After an initial questionnaire and baseline monitoring of adult Ae. aegypti abundance with BG-Sentinel (BGS) traps in six clusters, three clusters were randomly assigned to the intervention arm where each participating household received three MQTs for mass trapping during 17 months. The remaining three clusters (control arm) did not receive traps. The effect of mass trapping on adult Ae. aegypti abundance was monitored fortnightly with BGS traps. During the last two months of the study, a serological survey was conducted. After the study, a second questionnaire was applied in the intervention arm. Entomological monitoring indicated that MQT mass trapping did not reduce adult Ae. aegypti abundance. The serological survey indicated that recent dengue infections were equally frequent in the intervention and the control arm. Most participants responded positively to questions concerning user satisfaction. According to the results, there is no evidence that mass trapping with MQTs can be used as a part of dengue control programs. The use of this sticky trap is only recommendable for dengue vector monitoring.

Study of Vaccinia and Cowpox viruses' replication in Rac1-N17 dominant-negative cells

Interfering with cellular signal transduction pathways is a common strategy used by many viruses to create a propitious intracellular environment for an efficient replication. Our group has been studying cellular signalling pathways activated by the orthopoxviruses Vaccinia (VACV) and Cowpox (CPXV) and their significance to viral replication. In the present study our aim was to investigate whether the GTPase Rac1 was an upstream signal that led to the activation of MEK/ERK1/2, JNK1/2 or Akt pathways upon VACV or CPXV' infections. Therefore, we generated stable murine fibroblasts exhibiting negative dominance to Rac1-N17 to evaluate viral growth and the phosphorylation status of ERK1/2, JNK1/2 and Akt. Our results demonstrated that VACV replication, but not CPXV, was affected in dominant-negative (DN) Rac1-N17 cell lines in which viral yield was reduced in about 10-fold. Viral late gene expression, but not early, was also reduced. Furthermore, our data showed that Akt phosphorylation was diminished upon VACV infection in DN Rac1-N17 cells, suggesting that Rac1 participates in the phosphoinositide-3 kinase pathway leading to the activation of Akt. In conclusion, our results indicate that while Rac1 indeed plays a role in VACV biology, perhaps another GTPase may be involved in CPXV replication.

Role of IL-4 in an experimental model of encephalitis induced by intracranial inoculation of herpes simplex virus-1 (HSV-1) / Papel da IL-4 em modelo experimental de encefalite induzida pela inoculação intracraniana do herpes simplex vírus-1 (HSV-1)

Herpes simplex virus-1 (HSV-1) is a pathogen that may cause severe encephalitis in humans. In this study, we aimed to investigate the role of interleukin-4 (IL-4) in a model of HSV-1 brain infection. IL-4 knockout (IL-4-/-) and wild type (WT) C57BL/6 mice were inoculated with 10(4) plaque-forming units of HSV-1 by the intracranial route. Histopathologic analysis revealed a distinct profile of infiltrating cells at 3 days post-infection (dpi). Infected WT mice presented mononuclear inflammatory cells while IL-4-/- mice developed meningoencephalitis with predominance of neutrophils. IL-4-/- mice had diminished leukocyte adhesion at 3 dpi when compared to infected WT animals in intravital microscopy study. Conversely no differences were found in cerebral levels of CXCL1, CXCL9, CCL3, CCL5 and TNF-α between WT and IL-4-/- infected mice. IL-4 may play a role in the recruitment of cells into central nervous system in this acute model of severe encephalitis caused by HSV-1.

O vírus herpes simplex-1 (HSV-1) é um patógeno que pode causar encefalite grave em humanos. Neste estudo, buscamos investigar o papel da interleucina-4 (IL-4) no modelo de infecção intracerebral por HSV-1. Camundongos C57BL/6 selvagens (WT) e deficientes no gene IL-4 (IL-4-/-) foram inoculados com 10(4) unidades formadoras de placas de HSV-1 por via intracraniana. A análise histopatológica revelou um padrão distinto de infiltrado leucocitário. Camundongos WT infectados apresentaram infiltrado de células mononucleares, enquanto camundongos IL-4-/- desenvolveram meningoencefalite com predomínio de neutrófilos 3 dias pós-infecção (dpi). Animais IL-4-/- tiveram menor adesão de leucócitos 3 dpi quando comparados aos animais WT infectados à microscopia intravital. Em contrapartida, não foram encontradas diferenças nos níveis cerebrais de CXCL1, CXCL9, CCL3, CCL5 e TNF-α entre camundongos WT e IL-4-/- infectados. Esses resultados sugerem que IL-4 pode desempenhar um papel no recrutamento de células no sistema nervoso central neste modelo agudo de encefalite grave causada pelo HSV-1.

Cocirculation of two dengue virus serotypes in individual and pooled samples of Aedes aegypti and Aedes albopictus larvae / Cocirculação de dois sorotipos do vírus dengue em amostras individuais e pools de larvas de Aedes aegypti e Aedes albopictus

INTRODUCTION: To detect dengue virus, eggs of Aedes sp were collected in the city of Belo Horizonte, Brazil, in 2007. METHODS: Egg samples were subsequently hatched and the larvae were tested for the presence of dengue virus RNA by RT-PCR. RESULTS: Among the Aedes aegypti larvae samples, 163 (37.4 percent) out of 435 were positive, including 32 (10.9 percent) of 293 individual larvae samples concomitantly positive for two serotypes. CONCLUSIONS: Virological surveillance detecting coinfected vectors in the field could represent an important strategy for understanding the numerous factors involved in the transmission and clinical presentation of dengue.

INTRODUÇÃO: Para a detecção do vírus da dengue, ovos de Aedes sp foram coletados em Belo Horizonte, Brasil, em 2007. MÉTODOS: Amostras de ovos eclodiram e suas larvas foram testadas para a presença de RNA do vírus dengue por RT-PCR. RESULTADOS: Das amostras de larvas de Aedes aegypti, 163 (37,4 por cento) de 435 foram positivas, incluindo 32 (10,9 por cento) das 293 amostras individuais que foram concomitantemente positivas para dois sorotipos. CONCLUSÕES: A vigilância virológica de vetores no campo poderia representar uma estratégia importante para a compreensão dos diversos fatores envolvidos na transmissão e apresentação clínica da dengue.

Epidemiologia da poxvirose bovina no Estado do Espírito Santo, Brasil / Epidemiology bovine poxviruses in the Espírito Santo State, Brazil

O trabalho consistiu no estudo epidemiológico do surto de poxvirose bovina no Estado do Espírito Santo no período de 2002 a 2005. A coleta dos dados deu-se em 28 propriedades de gado bovino leiteiro de 08 municípios do Sul do Estado do Espírito Santo com casos de doenças pústulo-vesicular em bovinos, no período de agosto de 2002 a maio de 2005. Observou-se que a prevalência da doença no gado bovino leiteiro variou de 10,2% a 100% das propriedades com média de 52,7%. Através de isolamento viral e PCR do gene Timidina Quinase (TK), identificou-se o vírus do gênero Orthopoxvirus como o agente etiológico da poxvirose bovina. Em todas as propriedades constatou-se uma perda econômica, principalmente relacionada à queda na produção leiteira. A poxvirose bovina assumiu um caráter epidêmico no Sul do Estado do Espírito Santo, com grande impacto econômico e importância em saúde pública, como uma zoonose ocupacional entre os ordenhadores. Baseadas nos dados obtidos, medidas de cunho fiscal-sanitário e de orientação ao produtor agropecuário foram implementadas, visando conter a disseminação da doença entres as propriedades, e os outros municípios do Estado.

The purpose of this study was to investigate the epidemiological data of bovine poxviruses in the State of Espírito Santo during the years 2002 to 2005. The sample consisted of 28 properties that produce cow's milk in 08 cities from the South of the State, in wich were present cases of bonive pustulo-vesicular disease between 2002 august to 2005 may. The bovine disease was found in 10,2% to 100% of the properties with a mean of 52,7%. Throughout virus isolation techniques and PCR of the Timidin Kinase gene, an Orthopoxvirus was identified as the etiological agent of the bovine yiruses. All the properties reported economic losts related to a decrease in milk production. The bovine poxviruses assumed an epidemic feature in the South of the State of Espírito Santo promoting a big economic impact and representing an important public health worry as an occupational zoonosis mainly for the milkers. Thus, based on the obtained data, policy and sanitary statements, and educational strategies, were applied in the properties in order to stop the disease dissemination over the neighbor cities.

A rapid polymerase chain reaction protocol to detect adenovirus in eye swabs

Purpose: Viruses of the Adenoviridae family are associated with many clinical syndromes, processing 50 serotypes. These agents and viruses of the Herpesviridae family are the two major agents responsible for viral conjunctivitis, and a rapid diagnosis is important due to the epidemic character of adenoviral infection. Methods: We developed a PCR without DNA extraction for adenovirus using primers thath amplify a 300 tp gragament of the hexon capsid protein gene from many serotypes. Results: Swab samples from cornea of seven patients with keratoconjunctivitis were analyzed, and one of them was PCR positive for adenovirus. The sequence of this fragment shows a 100 percent homology with the sequence of adenovirus type 8. Conclusion: Sequencing of 300 bp from the hexon gene allows to identity almost all Ad serotypes, including all serotypes related to epidemic keratokonjuntivitis (8, 19,37) and almost all serotypes involved with Ad-associated conjunctivitis.

Genetic variability of HIV-1 isolates from Minas Gerais, Brazil

We report results of nucleotide sequencing and phylogenetic analysis of the env gene of 11 HIV-1 isolates, in Belo Horizonte, Brazil. Ten isolates belonged to HIV-1 subtype B and one was a probable B/F mosaic. This putative B/F recombinant is similar but not identical in its nucleotide sequence to other B/F mosaics described in Brazil.

Produçäo de interferon humano de membranas amnióticas em microtécnica / Production of human amniotic interferon by microtechnique

Foi desenvolvido um sistema de produçäo de interferon humano de membranas amnióticas (IFN-MA) em microtécnica. Fragmentos de âmnio de 1,4 ou 2,2cm de diâmetro, em placas de microtécnica (24 câmaras), foram infectados por vírus Sendai e o IFN-MA resultante foi titulado. Maiores títulos (12.800 e 13.000 unidades por ml) foram obtidos quando utilizadas quantidades de meio de 0,5ml e 1,0ml); 6,4 unidades hemaglutinantes do vírus Sendai e um fragmento por câmara. Níveis de IFN-MA mais consistentes e altos foram alcançados na regiäo coriônica do âmnio (6.000 a 9.600 unidades/ml) quando comparados com aqueles da regiäo umbilical (860 a 2.500 unidades/ml) e reflexa (200 a 6.500 unidades/ml). O sistema com fragmentos de 2,2cm de diâmetro produziu, em média, quantidades de interferon/ml superiores (9.300 unidades/ml) quando comparado ao da produçäo do âmnio total (2.200 unidades/ml). Apesar da variabilidade nos títulos produzidos individualmente pelos fragmentos de 2,2cm de diâmetro, devido a econômia de tempo e de materiais e aos altos níveis de IFN resultantes, a microtécnica poderá se empregada quando um grande número de variáveis for investigada