Comparative analysis of SARS-CoV-2 variants Alpha (B.1.1.7), Gamma (P.1), Zeta (P.2) and Delta (B.1.617.2) in Vero-E6 cells: ultrastructural characterization of cytopathology and replication kinetics

Abstract This study compares the effects of virus-cell interactions among SARS-CoV-2 variants of concern (VOCs) isolated in Brazil in 2021, hypothesizing a correlation between cellular alterations and mortality and between viral load and transmissibility. For this purpose, reference isolates of Alpha, Gamma, Zeta, and Delta variants were inoculated into monolayers of Vero-E6 cells. Viral RNA was quantified in cell supernatants by RT‒PCR, and infected cells were analyzed by Transmission Electron Microscopy (TEM) for qualitative and quantitative evaluation of cellular changes 24, 48, and 72 hours postinfection (hpi). Ultrastructural analyses showed that all variants of SARS-CoV-2 altered the structure and function of mitochondria, nucleus, and rough endoplasmic reticulum of cells. Monolayers infected with the Delta variant showed the highest number of modified cells and the greatest statistically significant differences compared to those of other variants. Viral particles were observed in the cytosol and the cell membrane in 100 % of the cells at 48 hpi. Alpha showed the highest mean particle diameter (79 nm), and Gamma and Delta were the smallest (75 nm). Alpha and Gamma had the highest particle frequency per field at 48 hpi, while the same was observed for Zeta and Delta at 72 hpi and 24 hpi, respectively. The cycle threshold of viral RNA varied among the target protein, VOC, and time of infection. The findings presented here demonstrate that all four VOCs evaluated caused ultrastructural changes in Vero-E6 cells, which were more prominent when infection occured with the Delta variant.

Monkeypox (Mpox) virus isolation and ultrastructural characterisation from a Brazilian human sample case

BACKGROUND According to the last 2023 Monkeypox (Mpox) Outbreak Global Map from the Centres for Disease Control and Prevention (CDC), more than 100 countries with no Mpox infection report cases. Brazil stands out in this group and is the second country with the highest number of cases in the last outbreak. OBJECTIVE To contribute to knowledge of the virus infection effects in a cellular model, which is important for diagnosis infections not yet included in a provider´s differential diagnosis and for developing viral inhibition strategies. METHODS We describe a virus isolation protocol for a human clinical sample from a patient from Brazil, the viral growth in a cell model through plaque forming units (PFU) assay, reverse transcriptase polymerase chain reaction (RT-PCR) and transmission electron microscopy (TEM). FINDINGS We follow the viral isolation in Vero cell culture from a Mpox positive clinically diagnosed sample and show the infection effects on cellular structures using a TEM. MAIN CONCLUSIONS Understanding the impact of viral growth on cellular structures and its replication kinetics may offer better strategies for the development of new drugs with antiviral properties.

Primary infection of BALB/c mice with a dengue virus type 4 strain leads to kidney injury

BACKGROUND Dengue is a disease caused by dengue virus (DENV-1 through -4). Among the four serotypes, DENV-4 remains the least studied. Acute kidney injury is a potential complication of dengue generally associated with severe dengue infection. OBJECTIVES The goal of this study was to investigate the alterations caused by experimental dengue infection in the kidney of adult BALB/c mice. METHODS In this study, BALB/c mice were infected through the intravenous route with a DENV-4 strain, isolated from a human patient. The kidneys of the mice were procured and subject to histopathological and ultrastructural analysis. FINDINGS The presence of the viral antigen was confirmed through immunohistochemistry. Analysis of tissue sections revealed the presence of inflammatory cell infiltrate throughout the parenchyma. Glomerular enlargement was a common find. Necrosis of tubular cells and haemorrhage were also observed. Analysis of the kidney on a transmission electron microscope allowed a closer look into the necrotic tubular cells, which presented nuclei with condensed chromatin, and loss of cytoplasm. MAIN CONCLUSIONS Even though the kidney is probably not a primary target of dengue infection in mice, the inoculation of the virus in the blood appears to damage the renal tissue through local inflammation.

Foraging preferences of the native stingless bee Melipona seminigra pernigra (Apidae: Meliponini) in campo rupestre on canga of Serra dos Carajás, southeastern Amazonia / Preferências de forrageamento da abelha nativa sem ferrão Melipona seminigra pernigra (Apidae: Meliponini) em campo rupestre na canga da Serra dos Carajás, sudeste da Amazônia

Abstract: Honey pollen samples of Melipona seminigra pernigraMoure & Kerr 1950 sampled between 2017 and 2019 from experimental apiaries installed in campo rupestre on canga (CRC) vegetation of the Serra dos Carajás aimed to evaluated seasonal floral availability of undisturbed and mining-influenced areas. Around one hundred pollen types were identified mainly belonging to Fabaceae, Myrtaceae and Euphorbiaceae (31, 6 and 5 species, respectively). Mining area presented the highest pollen richness, almost twice those identified in the undisturbed areas. 80% of the pollen types are rare with concentrations ≤ 2,000 pollen grains/10 g, while the remaining were the most abundant, frequent and the primary bee sources. These latter correspond mostly to native plants species such as Tapirira guianensis Aubl., Protium spp., Aparisthmium cordatum (A.Juss.) Baill., Mimosa acutistipula var. ferrea Barneby, Periandra mediterranea (Vell.) Taub., Miconia spp., Pleroma carajasense K.Rocha, Myrcia splendens (Sw.) DC., Serjania spp. and Solanum crinitum Lam. All pollen types were identified during both seasons, but higher concentration values are related to the dry period (June-September). The statistical analysis of the pollen data indicated that there was no significant difference between undisturbed and mining-influenced areas, since primary bee sources of this study are widespread used in revegetation of mined areas.

Resumo: O conteúdo polínico de amostras de mel coletadas nos anos de 2017 e 2019 de apiários experimentais de Melipona seminigra pernigraMoure & Kerr 1950, instalado dentro de uma vegetação de campo rupestre em um afloramento de canga na Serra dos Carajás, sudeste da Amazônia, foi analisado para entender a variabilidade local dos recursos florais em áreas naturais e perturbadas. Aproximadamente 100% dos tipos polínicos foram identificados e pertencem principalmente às famílias Fabaceae, Myrtaceae e Euphorbiaceae (31, 6 e 5 espécies, respectivamente). Áreas de mineração apresentaram a maior riqueza de pólen, quase o dobro daquelas identificadas em áreas perturbadas. 80% dos tipos de pólen são raros com concentrações ≤ 2.000 grãos de pólen/10g, enquanto que os restantes foram os mais abundantes, frequentes e fontes primárias para as abelhas. Este últimos correspondem principalmente a plantas nativas como Tapirira guianensis Aubl., Protium spp., Aparisthmium cordatum (A.Juss.) Baill., Mimosa acutistipula var. ferrea Barneby, Periandra mediterrânea (Vell.) Taub., Miconia spp., Pleroma carajasense K.Rocha, Myrcia splendens (Sw.) DC., Serjania spp. e Solanum crinitum Lam. Todos os tipos polínicos foram identificados durante ambas as estações, mas altas concentrações estão relacionadas ao período seco (junho-setembro). A análise estatística indicou que não houve diferença significativa nos dados de pólen de mel entre áreas naturais e áreas anteriormente degradadas, uma vez que as fontes primárias das abelhas deste estudo são amplamente utilizadas na revegetação de áreas mineradas.

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.

Low impact to fixed cell processing aiming transmission electron microscopy

In cell culture, cell structures suffer strong impact due to centrifugation during processing for electron microscope observation. In order to minimise this effect, a new protocol was successfully developed. Using conventional reagents and equipments, it took over one week, but cell compression was reduced to none or the lowest deformation possible.

Mycotoxicological and palynological profiles of commercial brands of dried bee pollen

Abstract Pollen is used in the human diet as a food supplement because of its high nutritional value; however, this product is prone to fungal contamination that could potentially generate toxins that are harmful to human health. This study aimed to verify the floral diversity of commercial brands of bee pollen and their mycotoxicological safety for human consumption. A total of 27 bee pollen samples were analyzed; these samples represented commercial brands, either showing an inspection seal or not, marketed in the State of Rio de Janeiro. The analyzed parameters included floral diversity through palynological analysis, water activity, fungal counts, identification and toxigenic profiles. The palynological analysis identified nine plant families, of which the Asteraceae was predominant. Analysis of hygienic quality based on fungal load showed that 92% of samples were reproved according to the commercial, sanitary, and food safety quality indicators. Aspergillus, Cladosporium and Penicillium were the most common genera. Toxigenic evaluation showed that 25% of the A. flavus strains produced aflatoxins. The high rate of contamination of products bearing an inspection seal emphasizes the need to monitor the entire procedure of bee pollen production, as well as to revise the current legislation to ensure safe commercialization of this product.

Bioterrorismo e microrganismos patogenicosApresentacao / Bioterrorism and pathogenic microorganisms

O uso de microrganismos patogênicos em atos de bioterrorismo é já há algum tempo objeto de grande preocupação em vários países. O presente trabalho apresenta a possível aplicação de vírus e bactérias para fins bélicos e terroristas, bem como o diagnóstico laboratorial para a identificação desses agentes. Foram salientados, entre outros, como agentes de infecções humanas visando o bioterrorismo, os vírus da varíola (ortopoxvírus), os de febres hemorrágicas e os pertencentes aos filovírus. Entre as bactérias foram destacadas as do antrax ( Bacillus anthracis ), da peste ( Yersinia pestis ), do botulismo ( Clostridium botulinum ) e da tularemia ( Francisella tularensis ), incluindo ainda a ricina ( Ricinus communis ) como componente do grupo B de agentes.

In recent years the use of pathogenic microorganisms in acts of bioterrorism has been the subject of major concern in many countries. This paper presents a possible application of viruses and bacteria for warfare and terrorist purposes, as well as a laboratory diagnosis to identify those agents. The viruses of smallpox (orthopoxvirus), of hemorrhagic fever and those belonging to filovirus have been highlighted, inter alia, as agents of human infection with bioterrorist intent. Among the bacteria, the emphasis has been on anthrax (Bacillus anthracis), the plague (Yersinia pestis), botulism (Clostridium botulinum) and tularemia (Francisella tularensis), not to mention ricin (Ricinus communis), as one of the Group B agents.

Floral sources to Tetragonisca angustula (Hymenoptera: Apidae) and their pollen morphology in a Southeastern Brazilian Atlantic Forest

The stingless bees are important flowers visitors of several plant species, due to their feeding habits and foraging behavior, constituting an important group to maintain biodiversity and the dynamics of tropical communities. Among stingless bees, Tetragonisca angustula is widely distributed in tropical habitats, and has been considered an important pollinator of different plant families. To support a rational economic use of this group, there is a need to characterize the plant species that represent important sources as part of their diet, as preferred, alternative or casual food sources. The aim of this survey was to distinguish the plant species that T. angustula visited most often. The study was undertaken in four regions of the Atlantic Rainforest in Rio de Janeiro state (Brazil) over a year from March 2008 to February 2009. For this, we collected bees, flowering plants and bee pollen loads from the four sites, and evaluated pollen morphology in the laboratory. Field observations showed the presence of plants belonging to ten different families and pollen loads showed the presence of pollen types belonging to 26 plant families. There were strong differences between pollen types, especially regarding pollen grain shape. The present survey suggests a high value of these plant species as trophic resources for the T. angustula in the understory of Atlantic Rainforest. Changes in these fragments of this forest may compromise the availability of resources for Tetragonisca angustula species and other stingless bees.

Para apoyar el uso racional de las abejas sin aguijón, es necesario conocer las especies de plantas que actúan como fuentes de recursos para estas abejas en su ambiente natural. El objetivo de este estudio fue identificar las especies de plantas que fueron visitadas con mayor frecuencia por abejas Tetragonisca angustula y describir los granos de polen de estas plantas. El estudio se realizó en la Mata Atlántica, donde se recogieron las abejas, las plantas con flores y el polen de las cargas corbiculares de las abejas obreras. La observación de campo mostró la presencia de plantas pertenecientes a diez familias y las cargas de polen mostraron la presencia de tipos de polen pertenecientes a 26 familias botánicas. Hubo grandes diferencias entre los tipos de polen, sobre todo teniendo en cuenta la ornamentación de los granos de polen. Este estudio sugiere un alto valor de estas especies de plantas como recursos tróficos para las abejas jataí en el sub-bosque de la Mata Atlántica. Las alteraciones de los fragmentos de bosque pueden afectar la disponibilidad de recursos para Tetragonisca angustula y otras abejas sin aguijón y ser un gran obstáculo para la su crianza sostenible.

Rudolf Barth: um cientista pioneiro na ilha da Trindade / Rudolf Barth: a pioneering scientist on the island of Trindade

Em consequência da instituição do Ano Geofísico Internacional, em 1957, a Marinha do Brasil organizou uma expedição à ilha da Trindade, da qual participou Rudolf Barth, pesquisador do Instituto Oswaldo Cruz e do Instituto de Pesquisas da Marinha, membro da Academia Brasileira de Ciências e do Conselho Nacional de Pesquisas. Esse cientista retornou à ilha da Trindade no ano seguinte, em uma segunda viagem com a Marinha brasileira. As pesquisas feitas então resultaram em algumas publicações científicas e em dois relatórios inéditos com observações e dados concernentes a fenômenos zoológicos (terrestres), meteorológicos, climatológicos, geomorfológicos e biológicos. Os relatórios são transcritos a seguir.

As a result of the institution of the International Geophysical Year in 1957, the Brazilian Navy organized an expedition to the island of Trindade, on which Rudolf Barth participated. He was a researcher at the Oswaldo Cruz Institute and the Naval Research Institute, and a member of the Brazilian Academy of Sciences and the National Research Council. The scientist returned to the island of Trindade the following year, on a second expedition with the Brazilian Navy. The research findings then resulted in some scientific publications and two unpublished reports with observations and data concerning terrestrial, zoological, meteorological, climatological, geomorphological and biological phenomena. The reports are then transcribed.

Conociendo la miel de Melipona favosa en la Península de Paraguaná, estado Falcón, Venezuela / Knowing the Melipona favosa honey from Paraguaná Peninsula, Falcon state, Venezuela

Diversas especies de abejas sin aguijón producen miel de botija en Venezuela, la cual no está incluida en las normas venezolanas de calidad. Una de estas especies, la Meli po na favosa, vive en la Península de Paraguaná, donde se conoce como erica o maba. A fin de conocer las mieles de M. favosa, se realizó la determinación de su origen botánico, su caracterización físicoquímica según los métodos de la norma COVENIN 2136-84 para miel de abejas y contenido de nitrógeno por microkjeldahl, y de actividad biológica con métodos espectrofotométricos y de concentración inhibitoria mínima, en seis mieles. Son mieles claras de color ámbar entre 18 y 79 mm Pfund. La composición físicoquímica varió así: acidez libre 12,72-95,86 meq/kg, pH 3,53-4,44, humedad 25,40-32,00 g agua/100 g, 0,01-0.16 g ce ni zas/100g, 10,48-57,55 mgN/100 g, 62,60-69,50 g azúcares reductores/100 g, 0,60-5,10 g sacarosa aparente/100 g, flavonoides 0,10-8,15 mgEQ/100g miel, y polifenoles 51,50-217,19 mgEAG/100 g. La actividad antibac teriana (g miel/100 mL medio de cultivo) fue mayor contra E. coli (12.50-50,00) que contra S. aureus (50,00- 50,00). La actividad antioxidante se ubicó en un rango bajo a alto para mieles, con 45,91-227,92 mmoles equivalentes de Trolox/100 g. El análisis melisopalinológico indicó polen dominante del género Portulaca y la especie Carica papaya.

Diverse species of stingless bees produce por honey in Venezuela,which is not included in the Venezuelan regulations of honey quality. One of these species , the Melipona favosa, lives in the Paraguaná Peninsula, where it is known as erica or maba. In order to know the honeys of M. favosa, the botanical origin was determined, their physicochemical characterization was done according to the methods of the norm COVENIN 2136-84 for honey, the nitrogen content by microkjeldahl, and biological activity by spectrophotometric and minimum inhibitory concentration methods, in six ho neys. They are light honey amber color between 18 and 79 mm Pfund. The physicochemical composition varied as follows: free acidity 12.72- 95.86 meq/kg, pH 3.53-4.44, moisture 25.40-32.00 g water/100 g, 0.01-0.16 g ash/100g, 10.48-57.55 mgN/100 g, 62.60-69.50 g reducing su gars/100 g, 0.60-5.10 g apparent sucrose/100 g, flavonoids 0.10-8.15 mgQE/100g honey, and polyphenols 51.50-217.19 mgGAE/100 g. The antibacterial ac tivity (g ho ney/100 mLculture media) was higher against E. coli (12.50-50.00) than S. aureus (50.00-50.00). The antioxidant activity was in a low to high range for honeys, with 45.91- 227.92 mmoles Trolox equivalents/100 g.

Infecções humanas causadas por poxvirus relacionados ao vírus vaccinia no Brasil / Human infections caused by vaccinia-like poxviruses in Brazil

A partir de 1999, infecções humanas por Orthopoxvirus vem sendo observadas em pelo menos oito estados no país, com a formação de vesículas as quais evoluem para pústulas e crostas, principalmente nos membros superiores e face, após contacto com bovinos apresentando lesões semelhantes no úbere. Alem das lesões na pele, foram descritas nos pacientes reações ganglionares axilares por vezes dolorosas, febre, cefaléia, fadiga, desidratação, anorexia, sudorese, artralgia e mialgia, evoluindo o quadro por três a quatro semanas. Lesão vulvar bem como transmissão intrafamiliar foram igualmente descritas. Estudos moleculares demonstraram que os poxvirus identificados são geneticamente relacionados a amostras do vírus vaccinia utilizadas no passado, nas campanhas de vacinação. Especimens clínicos de 80 infecções humanas foram estudados no laboratório e a infecção por orthopoxvirus confirmada em 68 casos. São apresentadas lesões observadas em pacientes bem como discutidas as implicações desta zoonose no Brasil.

Since 1999, human infection caused by Orthopoxvirus has been observed in at least eight Brazilian states, with the presence of vesicles that evolve to pustules and crusts, especially on the hands, arms and face, after contact with cows showing comparable lesions on the udder. In addition to the skin lesions, there have been descriptions of patients with axillary ganglionic reactions that are sometimes painful, along with fever, headache, fatigue, dehydration, anorexia, sudoresis, arthralgia and muscle pain. The condition evolves over a three to four-week period. Vulvar lesions and transmission within families have also been described. Molecular studies have shown that the poxviruses identified are genetically related to vaccinia virus samples that were used in vaccination campaigns in the past. Clinical specimens from 80 human infections were studied in the laboratory, and orthopoxvirus infections were confirmed in 68 cases. The lesions observed in these patients are presented and the implications of this zoonosis in Brazil are discussed.

Relação entre a composição nutricional e a origem floral de pólen apícola desidratado / Correlation between nutritional composition and floral origin of dried bee

A composição química do pólen é variável e depende da origem botânica. No presente trabalho foi investigada a correlação da composição centesimal e das vitaminas antioxidantes com a origem floral em amostras de pólen apícola. Foram analisadas seis amostras desidratadas de pólen apícola coletadas na região do Vale do Ribeira, São Paulo, Brasil. A vitamina C foi quantificada por titulação, o β-caroteno por cromatografia em coluna aberta e a vitamina E por cromatografia líquida de alta eficiência. A composição centesimal foi determinada seguindo-se as metodologias recomendadas por AOAC (1995) e IAL (2005) e a caracterização floral foi feita por meio de observação em microscópio. As concentrações das vitaminas variaram de 114 a 340μg/g para vitamina C, 16,27 a 38,64μg/g para vitamina E e de 3,14 a 77,88μg/g para o β-caroteno. A composição centesimal das amostras apresentou conformidade com as especificações estabelecidas pela legislação brasileira. Foi encontrada grande variabilidade de tipos polínicos nas amostras analisadas e alguns deles apresentaram alta correlação com a vitamina C (Myrtaceae), com o β-caroteno (Arecaceae, Cecropia e Fabaceae) e com os lipídeos (Arecaceae e Fabaceae). Outras amostras mostraram correlação negativa com β-caroteno (Mimosa caesalpiniaefolia e Poaceae), com proteínas(Arecaceae) e com lipídeos (Mimosa caesalpineafolia).

Tipos polínicos encontrados em colônias de abelhas africanizadas sujeitas à doença cria ensacada brasileira / Pollen types found in africanized honeybees colonies submitted to brazilian sac brood disease

O pólen de algumas espécies de Stryphnodendron (Mimosoidea), conhecidas popularmente por barbatimão, tem sido apontado como agente causador da mortandade de larvas pré-pupas observada na cria ensacada brasileira (CEB). Neste estudo, objetivou-se identificar os tipos polínicos encontrados em amostras de pólen coletadas em colméias de Apis mellifera L., no Estado do Rio de Janeiro, a fim de elucidar as fontes florais utilizadas por essa espécie de abelhas em períodos sujeitos a CEB, com especial atenção para a presença do pólen de Stryphnodendron spp. Foram coletadas amostras de pólen apícola e pão de abelhas em sete apiários, em três municípios do Estado do Rio de Janeiro. A amostragem foi tomada um mês antes e durante a ocorrência da CEB. As famílias botânicas mais representativas foram: Leguminosae, Euphorbiaceae, Asteraceae, Poaceae e Myrtaceae. O pólen de Stryphnodendron não esteve presente nas amostras analisadas. A ausência de Stryphnodendron no menu floral das colônias africanizadas acometidas pela CEB em condições naturais sugere outro fator causal para a CEB no Estado do Rio de Janeiro.

The pollen of some Stryphnodendron species (Mimosoidea), commonly known as barbatimão, has been identified as a causative agent of pre-pulps larvae mortality, observed in Brazilian sac brood disease (BSB). The goal of this study is to identify the pollen types found in samples of pollen collected by Apis mellifera L. to elucidate the floral diet used by this honeybee specie in Rio de Janeiro state during the periods where BSB usually occurs. Particular attention was given to the pollen of Stryphnodendron spp presence. Samples of pollen and bee bread were collected from seven apiaries located in Rio de Janeiro state, Brazil. Sampling procedures were done a month before BSB usual occurrence and also during its occurrence. Leguminosae, Euphorbiaceae, Asteraceae, Poaceae and Myrtaceae were most representative botanical families on pollen sampling. Pollen from Stryphnodendron was not presented at the analyzed samples. In order to Stryphnodendron absent at the floral diet of Africanized hives affected by BSB in natural conditions, it is proposed another causal factor for BSB in the state of Rio de Janeiro.

Histopathological and ultrastructural aspects of mice lungs experimentally infected with dengue virus serotype 2

Histological and ultrastructural alterations in lung tissue of BALB/c mice infected with dengue virus serotype 2 (non-neuroadapted), by intraperitoneal and intravenous routes were analyzed. Lung tissues were processed following the standard techniques for photonic and electron transmission microscopies. Histopathological and ultrastructural studies showed interstitial pneumonia, characterized by the presence of mononuclear cells. In the mouse model, the dengue virus serotype 2 seems to led to a transient inflammatory process without extensive damage to the interalveolar septa, but caused focal alterations of the blood-exchange barrier. Endothelial cells of blood capillaries exhibited phyllopodia suggesting activation by presence of dengue virus. Morphometrical analysis of mast cells showed an expressive increase of the number of these cells in peribronchiolar spaces and adjacent areas to the interalveolar septa. Alveolar macrophages showed particles dengue virus-like inside rough endoplasmic reticulum and Golgi complex, suggesting viral replication. The tissue alterations observed in our experimental model were similar to the observed in human cases of dengue fever and dengue hemorrhagic fever. Our results show that BALB/c mice are permissive host for dengue virus serotype 2 replication and therefore provides an useful model to study of morphological aspects of dengue virus infection.

Morphological studies in a model for dengue-2 virus infection in mice

One of the main difficulties in studying dengue virus infection in humans and in developing a vaccine is the absence of a suitable animal model which develops the full spectrum of dengue fever, dengue haemorrhagic fever, and dengue shock syndrome. It is our proposal to present morphological aspects of an animal model which shows many similarities with the dengue infection in humans. BALB/c mice were intraperitoneally infected with non-neuroadapted dengue virus serotype 2 (DENV-2). Histopathological and morphometrical analyses of liver tissue revealed focal alterations along the infection, reaching wide-ranging portal and centrolobular veins congestion and sinusoidal cell death. Additional ultrastructural observations demonstrated multifocal endothelial injury, platelet recruitment, and alterated hepatocytes. Dengue virus antigen was detected in hepatocytes and in the capillar endothelium of the central lobular vein area. Liver function tests showed high levels of aspartate transaminase and alanine transaminase enzyme activity. Lung tissue showed interstitial pneumonia and mononuclear cells, interseptal oedema, hyperplasia, and hypertrophy of the bronchiolar epithelial cells. DENV-2 led to a transient inflammatory process, but caused focal alterations of the blood-exchange barrier. Viremia was observed from 2nd to 11th day p.i. by isolation of DENV-2 in C6/36 mosquito cell line inoculated with the supernatant of macerated liver, lung, kidney, and cerebellum tissues of the infected mice.

In situ hybridization of hepatitis C virus RNA in liver cells of an experimentally infected rhesus macaque

The liver tissue of a rhesus macaque inoculated with hepatitis C virus (HCV) has been analyzed for the presence of HCV RNA using the technique of in situ hybridization, both at light and electron microscopy levels. The animal was inoculated by the intrasplenic route using a HCV infected autogenic hepatocyte transplant. The serum sample used to infect the hepatocyte cells was characterized by polymerase chain reaction technique and shown to be positive for HCV RNA, genotype 3 with 10(7) RNA copies/ml. In situ hybridization was performed using a complementary negative strand probe made with the specific primer. We were able to detect and localize viral RNA in altered membranes of the rough endoplasmic reticulum of infected liver cells, showing evidence of virus replication in vivo.

Detection of poxvirus in cattle associated with human cases in the State of Rio de Janeiro: preliminary report

This preliminary report describes human and cow cases of poxvirus that recently ocurred in the State of Rio de Janeiro. The electron microscopic findings were consistent with parapoxviral and orthopoxviral infection. Orthopoxvirus strains were isolated from human and cow cases. Detailed viral characterization by means of genetical techniques is under investigation. Based on these informations, poxviral diseases should be also considered an emerging viral zoonosis that can affect human beings.