Avaliação da infecção por Parvovírus Humano B19 em diferentes grupos populacionais: diagnóstico diferencial e aspectos imunopatogênicos da infecção aguda e persistente / Evaluation of Human Parvovirus B19 infection in different population groups: differential diagnosis and immunopathogenic aspects of acute and persistent infection

A infecção pelo Parvovírus B19 (B19V) pode ocorrer em indivíduos imunocompetentes e imunocomprometidos, de todas as faixas etárias, e se caracteriza por ser aguda e autolimitada, podendo levar a quadros de doença exantemática (DE), doença febril aguda (DFA), doença renal crônica (DRC) e falência hepática aguda (FHA). O diagnóstico diferencial de B19V nessas populações, muitas vezes, não ocorre e estudos sobre a prevalência do B19V são antigos e escassos, não refletindo a atualidade. Marcadores da infecção podem ser detectados na circulação e em diferentes tipos de tecidos, inclusive em tecidos não eritroides, por meses ou anos. A infecção pode levar a manifestações clínicas graves, que requer tratamento hospitalar, e a doenças inflamatórias atípicas, como: cardiomiopatia, artrite reumatoide, hepatite e vasculite. No entanto, a detecção de B19V DNA não implica necessariamente na presença de vírions infecciosos e na associação do B19V com essas manifestações atípicas. Dessa forma, o objetivo do trabalho foi otimizar técnicas de PCR em tempo real para quantificação do B19V DNA e de detecção de partículas virais infecciosas, a fim de realizar o diagnóstico diferencial da infecção pelo B19V em pacientes com DE, DFA, DRC e FHA. Para o diagnóstico da infecção, amostras de diferentes populações foram testadas: DE (n=54), DFA (n=60), DRC (n=221), e FHA (n=30). Amostras de soro (e de tecido hepático para FHA) foram submetidas a avaliação de marcadores sorológicos (IgM e IgG anti-B19V) e moleculares do B19V, a fim de determinar a fase da infecção em que o paciente se encontrava. Para a avaliação de marcadores moleculares, a metodologia de PCR quantitativo e em tempo real foi otimizada e permitiu um diagnóstico sensível e específico do B19V DNA. Além disso, a presença de vírions em amostras de pacientes com B19V (n=10) e de macacos cynomolgus (n=4) infectados experimentalmente foram avaliadas por meio da técnica de pré-tratamento das amostras com uma enzima endonuclease. O teste molecular (qPCR) otimizado durante o estudo, apresentou sensibilidade e especificidade de 100%. O ensaio com a endonuclease revelou que a maioria das amostras de soro humano tornou-se B19V DNA negativa após o pré-tratamento, indicando que não eram infecciosas. Foi observado prevalências do B19V DNA em 5,5% dos pacientes com DE; 6,6% em DFA; 65,6% em DRC, e 23,3% em FHA. Como conclusão a técnica de qPCR otimizada no presente estudo foi efetiva para o esclarecimento de casos da infecção por B19V e é adequada para diagnóstico diferencial. Além disso, o teste laboratorial baseado em endonuclease possibilitou a discriminação do B19V DNA (se encapsidado em vírions ou não). Portanto, estes testes podem ser utilizados para esclarecer o papel do B19V como agente etiológico associado a diversas manifestações clínicas. As prevalências encontradas nesse estudo indicam que o B19V está circulando entre os diversos grupos populacionais estudados e deve ser feita uma melhor vigilância da infecção, pois está presente tanto em indivíduos imunocompetentes como em imunocomprometidos. Além disso, os resultados sugerem a importância da inclusão de B19V no diagnóstico laboratorial diferencial, não apenas para fins epidemiológicos, mas também para o manejo adequado do paciente.

Parvovirus B19 (B19V) infection can occur in immunocompetent and immunocompromised individuals of all group ages and is characterized as acute and self limiting, which can lead to rash disease (RD), acute febrile illness (AFI), chronic kidney disease (CKD), and acute liver failure (ALF). Differential diagnosis of B19V in these populations often does not occur and studies on the prevalence of B19V are scarce, outdated, and do not reflect the current situation. B19V markers of acute infection can be detected in the circulation and in different tissue types, including non-erythroid tissues, for months to years and may lead to severe clinical manifestations, requiring hospital treatment, and to atypical inflammatory diseases, such as cardiomyopathy, rheumatoid arthritis, hepatitis, and vasculitis. However, the detection of B19V DNA does not necessarily imply the presence of infectious virions and the causal relation between B19V and atypical manifestations could not be proved yet. Thus, the aim of this study was to standardize the real-time PCR for quantification of B19V DNA and detection of infectious viral particles in order to perform the differential diagnosis of the B19V infection in RD, AFI, CKD, and ALF patients. For the diagnosis of the infection, samples from different populations were tested: RD (n=54), AFI (n=60), CKD (n=221), and ALF (n=30). Serum samples (and hepatic tissue for ALF) were submitted to the evaluation of B19V serological status (anti-B19V IgM and IgG antibodies) and molecular markers, in order to determine the stage of infection in which the patient is. For the evaluation of molecular markers, a quantitative real-time PCR methodology was optimized and allowed a sensitive and specific diagnosis of B19V DNA. In addition, the presence of virions in samples from patients with B19V (n=10) and from cynomolgus monkeys (n=4) experimentally infected were evaluated by endonuclease enzyme pretreatment. The molecular test optimized during the study showed 100% sensitivity and specificity. The endonuclease treatment assay revealed that most human serum samples became negative after pretreatment, as indicative of non-infective particles. Concerning the prevalence of B19V DNA: 5.5% were obtained in patients with RD; 6.6% in AFI; 65.6% in CKD, and 23.3% in ALF. In conclusion, the qPCR technique optimized in the present study was effective for clarifying cases of B19V infection and is suitable for differential diagnosis. In addition, the endonuclease-based laboratory test made it possible to discriminate B19V DNA (whether encapsidated in virions or not). Therefore, these tests can be used to clarify the role of B19V as an etiologic agent associated with several clinical manifestations. The prevalence found in this study indicate that B19V is circulating among the different populational groups that have been studied and better surveillance of the infection should be carried out, as it is present in both immunocompetent and immunocompromised individuals. In addition, the results suggest the importance of including B19V in the differential laboratory diagnosis, not only for epidemiological purposes but also for the proper management of the patient.

JMT-1: a novel, spherical lytic halotolerant phage isolated from Yuncheng saline lake

ABSTRACT This work described a novel halotolerant phage, JMT-1, with a spherical morphology. JMT-1, which was isolated from a hypersaline lake, could produce clear plaques on Chromohalobacter sp. LY7-3. The purified virions are spherical, have no visible tail, and are about 30-50 nm in diameter. JMT-1 has a wide host range, and this study showed that the phage can infect at least five halophilic bacteria. The proteins of JMT-1 were analyzed using sodium dodecyl sulfate polyacrylamide gel electrophoresis, and six proteins were detected. Results show that JMT-1 is a bacteriophage with a linear double-stranded DNA. Meanwhile, the genome is approximately 23 kb in length and is sensitive to the restriction endonucleases Bam I, EcoR I, Hind III and Kpa I. JMT-1 has a high titer, approaching 1.5 × 109 pfu/mL after dilution to 10−6 pfu/mL. The phage is also sensitive to chloroform but not to temperature, pH, and lowered salt concentration. JMT-1 is a spherical lytic halotolerant phage with a wide host range and has the tolerance to specific extreme environments. These data could provide references for studying phage resources in extreme environments and would also provide the useful methods for isolation and identification of other valuable phage in the salt lake environment.

Actividad biológica de tres Curcuminoides de Curcuma longa L. (Cúrcuma) cultivada en el Quindío-Colombia / Biological activity of three curcuminoids from Curcuma longa L. (turmeric) grown in Quindío, Colombia

Introducción: Curcuma longa L. es una planta de la familia Zingiberaceae distribuida en las regiones tropicales y subtropicales, utilizada en la industria alimentaria, en medicina y en cosmética. Su colorante principal es la curcumina, un polifenol con múltiples efectos medicinales. Objetivos: obtener, caracterizar químicamente y evaluar la actividad biológica de tres curcuminoides de C. longa, cultivada en el Quindío-Colombia. Métodos: se purificaron tres curcuminoides (curcumina (C), demetoxicurcumina (DMC) y bisdemetoxicurcumina (BDMC) desde el rizoma de la planta, en estado seco, por cromatografía en columna y se caracterizaron por punto de fusión, espectroscopía infrarroja (IR), espectroscopía UV-vis y espectrometría de masas. Se evaluó la actividad antimicrobiana en bacterias y hongos por el método modificado de pozos de agar, la citotoxicidad sobre células BHK-21 por el método de bromuro de 3-(4,5- dimetiltiazol-2-ilo)-2,5-difeniltetrazol (MTT) y la toxicidad sobre Artemia salina. Finalmente se determinó el efecto de los curcuminoides en células BHK-21 infectadas con dengue virus 2. Resultados: la curcumina presentó mayor punto de fusión (177,3 ºC-183,2 ºC). El espectro IR reveló los grupos funcionales característicos y el UV-vis indicó máximos de absorción para curcumina, demetoxicurcumina y bisdemetoxicurcumina de 419, 418 y 414 nm en cloroformo, respectivamente. El espectro de masas mostró m/z para C: 368, DMC: 338 y BDMC: 308. Se encontró actividad antimicrobiana frente a Staphylococcus aureus y Staphylococcus epidermidis, se determinó que BDMC presentó menor toxicidad y se evidenció mayor efecto inhibitorio sobre viriones infectivos de dengue con curcumina a 20 y 30 M. Conclusiones: la caracterización de los compuestos confirma su composición como polifenoles, lo cual se relaciona a la actividad biológica de éstos, encontrándose principalmente que la curcumina altera la infección por virus dengue en cultivo celular. Esta investigación confirma la importancia de los principios activos de plantas con amplio espectro farmacológico como C. longa(AU)

Introduction: Curcuma longa L. is a plant from the family Zingiberaceae distributed in tropical and subtropical regions and used in the food industry, in medicine and in cosmetics. Its main coloring substance is curcumin, a polyphenol with many medicinal properties. Objectives: Obtain, characterize chemically and evaluate the biological activity of three curcuminoids from C. longa grown in Quindío, Colombia. Methods: Three curcuminoids (curcumin (C), demethoxycurcumin (DMC) and bisdemethoxycurcumin BDMC) from the rhizome of the plant were purified in a dry state by column chromatography and characterized by fusion point, infrared (IR) spectroscopy, UV-vis spectroscopy and mass spectrometry. Antimicrobial activity against bacteria and fungi was evaluated by the modified agar well method, cytotoxicity to BHK-21 cells by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) method, and toxicity against Artemia salina. Finally, determination was made of the effect of the curcuminoids in BHK-21 cells infected with dengue virus 2. Results: Curcumin had the highest fusion point (177.3 ºC-183.2 ºC). IR spectroscopy revealed the characteristic functional groups and UV-vis spectroscopy showed maximum absorption values for curcumin, demethoxycurcumin and bisdemethoxycurcumin of 419, 418 and 414 nm in chloroform, respectively. Mass spectrometry found that m/z values were C: 368, DMC: 338 and BDMC: 308. Antimicrobial activity was observed against Staphylococcus aureus and Staphylococcus epidermidis. BDMC was found to have lower toxicity. A greater inhibitory effect against infective dengue virions was observed with curcumin at 20 y 30 µM. Conclusions: Characterization of the compounds confirms their polyphenolic composition, which manifests in their biological activity, mainly the capacity of curcumin to alter infection by dengue virus in cell cultures. The study corroborated the importance of the active principles of plants with a wide pharmacological spectrum, such as C. longa(AU)

Ultrastructure of Zika virus particles in cell cultures

Zika virus (ZIKV) has infected thousands of Brazilian people and spread to other American countries since 2015. The introduction of ZIKV brought a strong impact to public health in Brazil. It is of utmost importance to identify a susceptible cell line that will enable the isolation and identification of the virus from patient samples, viral mass production, and testing of drug and vaccine candidates. Besides real-time reverse transcriptase polymerase chain reaction diagnosis for detecting the viral genome, virus isolation in cell lines was useful in order to study the structure of the viral particle and its behaviour inside cells. Analysis of ZIKV infected cell lines was achieved using transmission electron microscopy (TEM). Blood was obtained from a Brazilian patient during the first days after presenting with signs of the disease, and ZIKV from the patient’s blood was isolated in the C6/36 mosquito cell line. Afterwards, Vero cells were inoculated with the viral suspension, fixed six days after inoculation, embedded in polymers, and ultra-thin cut. Like dengue viruses, this flavivirus showed numerous virus particles present inside cellular vesicles thereby confirming the susceptibility of the Vero cell line to ZIKV replication. TEM is a unique technique available to make the virus visible.

Bovine Herpesvirus 4 in Parana State, Brazil: case report, viral isolation, and molecular identification

Bovine Herpesvirus 4 (BoHV-4) is a member of Gammaherpesvirinae sub-family and belongs to genus Rhadinovirus. This virus has been associated with different clinical manifestations and research activity has put forward a strong correlation among virus infection, postpartum metritis, and abortion. The goal of this work was to characterize a virus strain isolate from a cow’s uterine outflow. From swabs drawn of uterine secretion, a virus strain was isolated and characterized by its cytopathology, morphology, and molecular biology approaches. In culture there was CPE development, characterized mainly by long strands with several small balloons along them, radiated from infected cells. Electron microscopy analysis revealed virus particles that had icosahedrical capsid symmetry surrounded by a loose envelope, typical of a herpesvirus. A 2,571 bp PCR product after HindIII digestion generated four fragments, whose base pair composition were 403, 420, 535, and 1,125 bp. Restriction enzymes HindIII and BamHI generated the expected diagnostic bands as well as a 2,350 bp hypermolar fragment as a result of BamHI treatment to demonstrate that agent was a bovine herpesvirus 4, appertaining to DN-599 group.

An improved purification procedure for Leishmania RNA virus (LRV)

Leishmania RNA Virus (LRV, Totiviridae) infect Leishmania cells and subvert mice immune response, probably promoting parasite persistence, suggesting significant roles for LRV in host-parasite interaction. Here we describe a new LRV1-4 purification protocol, enabling capsid visualization by negatively stained electron microscopy representing a significant contribution to future LRV investigations.

Expresión y purificación de las proteínas estructurales del rotavirus VP5* y VP8* en bacterias E. coli BL21(DE3) / Expression and purification of rotavirus structural proteins VP5* and VP8* in bacteria E. coli BL21(DE3)

La caracterización de las proteínas estructurales del rotavirus y de las proteínas de la superficie de la célula hospedera implicadas en la unión y penetración del virion requiere de la disponibilidad de cantidades suficientes y de alto grado de pureza de estas proteínas. Por lo tanto, el objetivo de este trabajo fue expresar y purificar las proteínas estructurales del rotavirus de la cepa RRV, VP5* y VP8*, y producir anticuerpos policlonales dirigidos contra ellas. Se expresaron las proteínas recombinantes VP5* (rVP5*) y VP8* (rVP8*) en bacterias E. coli BL21(DE3) transfectadas con el plásmido pGEX-4T que contenía sus secuencias codificantes. Se consideraron como variables el medio de crecimiento, número de bacterias antes de inducir la expresión, concentración del inductor y tiempo de inducción. La mayor proporción de rVP8* se obtuvo cuando las bacterias transformadas se cultivaron en medio LB y la inducción se llevó a cabo con 1 mM de IPTG cuando el cultivo alcanzó una OD 600 nm de 0.5 y la inducción se mantuvo durante 6 h. rVP5* alcanzó la mayor proporción cuando células a una OD 600 nm de 0.2 fueron inducidas con 0.5 mM de IPTG durante 4 h en medio 2XYT en presencia de glucosa al 2 %. Las proteínas recombinantes obtenidas, acumuladas en la fracción insoluble, fueron solubilizadas con detergentes iónicos y no iónicos, seguido de purificación mediante cromatografía de afinidad antes de ser empleadas como antígenos para la producción de anticuerpos policlonales en conejos. Estos anticuerpos se caracterizaron mediante su capacidad de reconocimiento de los antígenos correspondientes en ELISA, "Western blotting", y en ensayos de inmunocitoquímica en células infectadas con rotavirus RRV. La cantidad y el grado de pureza de las proteínas recombinantes obtenidas, y los anticuerpos dirigidos contra ellas, anticipan su utilidad como herramientas en la caracterización de la interacción virus-célula.

The characterization of rotavirus structural proteins and the cell surface proteins involved in virion binding and penetration depends on the availability of substantial amounts of highly purified proteins. The aim of the present work was to express and purify the rotavirus structural proteins VP5* and VP8* in order to produce polyclonal antibodies against them. Recombinant proteins VP5* (rVP5*) and VP8* (rVP8*) were expressed in E. coli cells transfected with pGEX-4T or pET 28a containing their corresponding encoding sequences. Culture medium, bacterial concentration before induction, inductor concentration and induction time were used as variables. The greater proportion of rVP8* was obtained when transfected bacteria were grown in medium LB and induction was started by adding 1 mM IPTG to cells at OD 600 nm 0.5 followed by 6 h-induction. The highest proportion of rVP5* was reached when cells at OD 600 nm 0.2 were induced with 0.5 mM IPTG for 4 h in medium 2XYT containing 2% glucose. The recombinant proteins accumulated in the insoluble fraction were solubilized with ionic and non-ionic detergents, and purified by affinity chromatography before being used as antigens for production of rabbit polyclonal antibodies. The antibodies produced were characterized through their ability to recognize the corresponding antigens in ELISA, Western blotting, and immunochemistry assays in rotavirus infected cells. The amount and purity of recombinant proteins obtained in this work, and the antibodies against them, are expected to be useful for the characterization of the virus-cell interaction.

Virioplankton abundance in trophic gradients of an upwelling field

This work correlates time series of biological and physical variables to the marine viruses across trophic gradients within Arraial do Cabo upwelling system, Southeast of Brazil. The objective is to investigate the major controlling factors of virioplankton dynamics among different water masses. It was used an in situ and ex situ flow cytometry for accessing the plankton community. Viruses were highly correlated to bacteria and phytoplankton, but although the lack of direct correlation with physicals, upwelling turned out to be the main contributing factor to the highest values of viral abundance and virus:bacterial ratio. Our data suggest that the lowest temperature of upwelled South Atlantic Central Waters would help to maintain a high viral abundance and higher temperatures of Coastal and Tropical Waters might be another ecological niche allowing the co-existence.

Mycobacteriophages as versatile tools for genetic manipulation of mycobacteria and development of simple methods for diagnosis of mycobacterial diseases / Micobacteriófagos como herramientas versátiles para la manipulación genética y el desarrollo de métodos simples para el diagnóstico de enfermedades micobacterianas

Tuberculosis, caused by Mycobacterium tuberculosis, is responsible for over two million deaths per year worldwide. Due to its long doubling time (18 h), the microbiological detection of M. tuberculosis by conventional methods takes up to one month, unless the number of bacilli in the biological sample is high enough. Thus, drug resistance assessment requires at least one month for obtaining the primary culture and another month to determine its susceptibility to antimycobacterial drugs. Moreover, for a long time, the lack of genetic tools for mycobacteria has been a barrier for undertaking studies aimed at understanding the mechanisms of drug resistance and drug target identification, being all these topics of utmost importance considering the increase in the number of drug-resistant clones and the few therapeutic options available. Mycobacteriophages are promising as a novel source of genetic elements for mycobacteria manipulation, as well as for the development of versatile, simple, fast and cheap methods for drug resistance assessment of M. tuberculosis clinical isolates. We herein describe the background related to the use of mycobacteriophages, with emphasis placed on their utilization for drug resistance analysis in our country.

La tuberculosis, enfermedad causada por el bacilo Mycobacterium tuberculosis, es responsable de más de dos millones de muertes anuales en el mundo. Debido a su largo tiempo de duplicación (18 h), la detección bacteriológica de M. tuberculosis por métodos convencionales necesita de un mes o aun más, a menos que el número de bacilos en la muestra clínica sea suficientemente alto. Por consiguiente, se necesita un mínimo de dos meses para determinar la resistencia de este microorganismo a las drogas antituberculosas: uno para obtener el cultivo primario y otro para ensayar la sensibilidad frente a aquellas. La falta de herramientas para la manipulación genética de micobacterias ha dificultado la identificación de los blancos de acción de las drogas y el estudio de los mecanismos de resistencia a éstas, tópicos de la mayor relevancia dado el aumento mundial del número de aislamientos clínicos multirresistentes y las pocas opciones terapéuticas disponibles. Los micobacteriófagos son considerados nuevas herramientas para la manipulación de las micobacterias, así como para el desarrollo de métodos simples, rápidos y económicos para determinar la sensibilidad a drogas de los aislamientos clínicos de M. tuberculosis. En esta revisión se describen los antecedentes del uso de micobacteriófagos con énfasis en su utilización para el análisis de resistencia a drogas antituberculosas en nuestro país.

An epitope variable-reservoir model for HIV-1 infection

Se presenta un modelo de infección por VIH-1 basado en el concepto de inmunodominancia. Además de viriones mutantes y células T-CD4, se considera también reservorios virales, entre ellos macrófagos y células dendríticas foliculares. También se considera ataque citotóxico contra los reservorios y ataque extracelular sobre los viriones, ambos coordinados por las células T-CD4. En primer lugar, se trabaja sólo con una variable viral, y se usan ciertas aproximaciones para obtener un modelo fácilmente manipulable, para el cual se encuentra un criterio de estabilidad que rige el paso de progresión a regresión de la infección. Este criterio mantiene su validez para el modelo sin uso de aproximaciones, esto es, dos epítopes que mutan al azar, cada uno de ellos con dos variantes. Los resultados sugieren: a) que el papel jugado por los reservorios en el mantenimiento de la infección es muy importante, b) que mantener el ataque de las células citotóxicas sobre los reservorios infectados facilita la labor del sistema inmune, y c) que la terapia debería orientarse preferiblemente hacia estos objetivos.

Citomegalovirus em abortamento humano: diagnóstico de processo infeccioso por sorologia e detecção direta de antígeno e de ácido nucleico virais / Citomegalovirus in human abortion: diagnosis of infectious process for sorologia and direct detention of antigen and acid nucléico you capsize

A possível correlação da infecção pelo HCMV em mulheres na menácme com perda reprodutiva até a 22ªsemana gestacional foi estudada neste trabalho.A população estudada foi constituída por mulheres de baixo nível sócio-econômico:em abortamento(89)e grupo controle (gestantes,40)e não gestantes(62).Espécimes clínicos obtidos foram:tecidos de abortamento(decídua,vilosidades coriônicas e fígado fetal,quando disponível),sangue,para obtenção de soro e de leucócitos(mulheres em abortamento e grupo controle)e secreção vaginal(grupo controle).A soroprevalência para HCMV foi de 97,8por cento.Anticorpos IgM foram detectados em quatro delas,2 em cada grupo,mas com IgG concomitantemente.A pesquisa do antígeno pp65 do HCMV,foi negativa em todas as amostras analisadas;por outro lado,o HCMV-DNA foi detectado em leucócitos de sangue periférico pelos métodos de PCR/nested-PCR em 7,1por cento,tanto das pacientes em abortamento quanto no grupo controle(5,1por cento nas gestantes e 8,5por cento nas não gestantes).A excreção cervical de HCMV,determinada pelos métodos de PCR/nested-PCR em secreção vaginal foi de 10por cento nas gestantes e de 14,5por cento nas não gestantes.Embora não tenha significado estatístico,a taxa de deteção menor em gestantes que em não gestantes sugere efeito supressivo viral no início da gestação.Todos os casos HCMV-DNA positivos a partir de secreção vaginal ou de leucócitos foram soropositivos para o HCMV mas IgM negativos.Dos 172 fragmentos de tecidos de abortamento(decídua,vilosidade coriônica e fígado fetal,quando disponível)obtidos de 89 casos,6,7por cento foram positivos pela PCR/nested-PCR. Vinte e três estirpes de HCMV(12 de secreção vaginal, três de leucócitos e oito de tecidos)obtidas pela PCR/nested-PCR do gene gB,foram genotipadas após digestão com enzimas de restrição(RFLP).O genótipo gB1 foi encontrado em freqüência maior(39,1por cento),seguido por gB2(26,1por cento)e por gB3 e gB4(17,4por cento,cada).Enquanto somente o genótipo gB1 foi encontrado em leucócitos,os 4 genótipos foram encontrados tanto em secreção vaginal quanto em tecidos de abortamento,sugerindo não existir um tropismo diferenciado para este tecido.O uso de sondas biotiniladas de HCMV,para localização tecidual da infecção do HCMV por hibridização in situ(HIS),não foi possível devido ao acúmulo de biotina endógena em núcleo de células do epitélio glandular da decídua em 10 de 11 casos estudados.A marcação das sondas com digoxigenina (DIG-11-dUTP)...

Biología de los virus de hepatitis / Biology of the hepatitis viruses

El alfabeto que conforman los virus causantes de hepatitis ha crecido en forma significativa en estos últimos años. Estos virus pertenecen a familias virales muy distintas y se dividen además en virus de transmisión entérica y transmisión parenteral. Antes de 1989, se conocía el VHA, picornavirus de transmisión entérica y sin mayores secuelas, el VHB, hepatitis de transmisión parenteral, con secuelas de cirrocis y cáncer de hígado y el VHD, daltavirus que requiere una co-infección por el VHB para poder replicarse en forma efectiva y responsable de muchos casos de hepatitis fulminantes. En 1989, se descubre por técnicas de biología molecular, el VHC, hepacivirus miembro de la familia flaviviridae, de transmisión parenteral cuya infección está asociada a secuelas similares a las del VHB. En 1990 se identifica el VHE, virus de transmisión entérica. El VHF es un término reservado a un virus de transmisión entérica, cuya identificación en la India es controversial. En la búsqueda del virus reponsable de la hepatitis post-transfusional no A hasta no E, entre 1995 y 1996 se describe el denominado VHG, de transmisión parenteral, utilizando herramientas cada vez más novedosas de biología molecular, aunque este virus no parece causar hepatitis. Mas recientemente se identifica el TTV(1997) y el Sen-V(2000), de nuevo por técnicas de biología molecular ¿Será algunos de estos virus al fin el responsable de las hepatitis post-transfusionales no A hasta no G?

Chimeric virus-like particles of the human papillomavirus type 16 (HVP 16) as a prophylactic and therapeutic vaccine

Infection by certain human papillomaviruses (HPV), most notably HPV types 16 and 18, is the mejor risk factor cervical cancer. Worldwide, this disease represents the second most frequent malignant tumor in women; thus, there is urgent need for efficient therapy and prevention. The natural history of cervical cancer and its precursors (cervical intraepithelial neoplasias), as well as animal experiments, strogly suggest that immune system controls both the primary infection (by neutralizing antibodies directed against the major structural protein L1) and the progression o the disease (via cytotoxic T cell specific for the viral oncoproteins expressed in transformed cells, e.g., E7). By the expression of an HPV 16 L1E7 fusion proteins, we have generated chimeric virus-like particles (CVLP). Immunization of mice with CVLPs induces neutralizing antibodies directed against L1 virus-like particles (devoid of the E7 portion) and E7-specific t cells as measured in vitro. Vaccinated animals are protected against tumor growth followings inoculation of syngeneic HPV 16-transformed cells. In addition, we observed a therapeutic effect of vaccination on pre-existing tumors. This data allowed us to concelude that CVLPs are suitable for prevention and therapy of HPV infection. A vaccine based on HPV 16 L1E7 CVLPs is currently under development

La dinámica del HIV y la restauración inmune: algunos avances y un gran desafío / The HIVïs dynamics and the immune restoration: some advances and a great challenge

Algunos descubrimientos recientes han permitido un mejor conocimiento de la fisiopatología de la enfermedad debida al HIV, lo que ha producido un cambio de las teorías hasta ahora aceptadas acerca de la replicación viral. De acuerdo con un modelo matemático recientemente desarrollado, la replicación viral alcanza proporciones astronómicas, lo que acompañado a numerosos errores que ocurren durante la transcripción del ARN permite la aparición de nuevas cuasi-especies. Las cepas mutantes son responsables en parte del fracaso de la monoterapia, hecho que es superado por la administración de tratamiento combinado que disminuye la replicación viral. Estos nuevos agentes terapéuticos han logrado aumentar el número de linfocitos TCD4(+) y reducir los niveles de la carga viral. Recientes publicaciones muestran que un pequeño grupo de pacientes presentaron infecciones oportunistas con un recuento de CD4 alto a pesar de estar recibiendo Terapéutica Antiviral Altamente Efectiva (HAART sus siglas en inglés). Por otro lado, algunos enfermos resolvieron sus patologías oportunistas a pesar de no recibir tratamiento específico contra las mismas, excepto HAART. Estos conceptos, y también otros referidos a la dinámica viral y la restauración inmune, son analizados en la presente revisión

Preliminar characterization of a new virus isolated from the spinal fluid of patients with meningitis in Säo Paulo, Brazil

De dezembro de 1982 a março de 1983 um total de 138 pacientes com idade de 4 meses a 57 anos foram atendidos em diferentes hosptais de Säo Paulo com sintomas de meningite asséptica. Um agente transmissível foi isolado de 35 das 53 amostras de líquor. A replicaçäo desse novo vírus ou de um grupo de vírus relacioandos antigenicamente com características semelhantes foi detectada através do efeito citopático produzido em cultura de células de MDCK. O agente isolado demonstrou ter um diâmtro em torno de 40 nm quando examinado ao microscópio eletrônico através da coloraçäo negativa. Nenhuma atividade hemaglutinante foi detectada em pH 7,2 com hemácias humanas de cobaias ou de galinha e em pH 6,0 - 7,2 com hemácias de ganso. Esse agente näo se mostrou patogênico ao camundongo recém-nascido e adulto, era sensível ao clorofórmio e näo foi inibido pelo BuDR, o que indica conter o genoma RNA

Cytoplasmic inclusions in virus-infected salivary gland cells of Triatoma infestans Klug.

Varios tipos de inclusoes citoplasmaticas (C1aC5) foram encontrados ao nivel da microscopia eletronica nas glandulas salivares de Triatoma infestans Klug infectadas con particulas viricas de cerca de 38-40nm e compostas de RNP. Admite-se que essas inclusoes sejam induzidas especificamente pela infeccao viral em consideracao.As inclusoes C1 a C4 parecem se tratar de etapas do processo de maturacao dos virions, enquanto as C5 podem ser remanescentes das inclusoes C4 ou resultar de corte relativamente superficial das ultimas. As inclusoes citoplasmaticas descritas nunca foram observadas no lumen das glandulas, mesmo durante a atividade secretora, embora particulas viricas livres sejam encontradas na secrecao salivar