Human virome in nasopharynx and tracheal secretion samples.

BACKGROUND: In Brazil the implementation of the Sentinel Surveillance System of Influenza began in 2000. Central public health laboratories use reverse transcription-quantitative polymerase chain reaction (RT-qPCR) for diagnosis of respiratory viruses, but this protocol identifies only specific targets, resulted in inconclusive diagnosis for many samples. Thus, high-throughput sequencing (HTS) would be complementary method in the identification of pathogens in inconclusive samples for RT-qPCR or other specific detection protocols. OBJECTIVES: This study aimed to detect unidentified viruses using HTS approach in negative samples of nasopharynx/tracheal secretions by the standard RT-qPCR collected in the Federal District, Brazil. METHODS: Nucleic acids were extracted from samples collected in winter period of 2016 and subjected to HTS. The results were confirmed by the multiplex PR21 RT-qPCR, which identifies 21 respiratory pathogens. FINDINGS: The main viruses identified by HTS were of families Herpesviridae, Coronaviridae, Parvoviridae and Picornaviridae, with the emphasis on rhinoviruses. The presence of respiratory viruses in the samples was confirmed by the PR21 multiplex RT-qPCR. Coronavirus, enterovirus, bocavirus and rhinovirus were found by multiplex RT-qPCR as well as by HTS analyses. MAIN CONCLUSIONS: Wide virus diversity was found by different methodologies and high frequency of rhinovirus occurrence was confirmed in population in winter, showing its relevance for public health.

Human virome in nasopharynx and tracheal secretion samples

BACKGROUND In Brazil the implementation of the Sentinel Surveillance System of Influenza began in 2000. Central public health laboratories use reverse transcription-quantitative polymerase chain reaction (RT-qPCR) for diagnosis of respiratory viruses, but this protocol identifies only specific targets, resulted in inconclusive diagnosis for many samples. Thus, high-throughput sequencing (HTS) would be complementary method in the identification of pathogens in inconclusive samples for RT-qPCR or other specific detection protocols. OBJECTIVES This study aimed to detect unidentified viruses using HTS approach in negative samples of nasopharynx/tracheal secretions by the standard RT-qPCR collected in the Federal District, Brazil. METHODS Nucleic acids were extracted from samples collected in winter period of 2016 and subjected to HTS. The results were confirmed by the multiplex PR21 RT-qPCR, which identifies 21 respiratory pathogens. FINDINGS The main viruses identified by HTS were of families Herpesviridae, Coronaviridae, Parvoviridae and Picornaviridae, with the emphasis on rhinoviruses. The presence of respiratory viruses in the samples was confirmed by the PR21 multiplex RT-qPCR. Coronavirus, enterovirus, bocavirus and rhinovirus were found by multiplex RT-qPCR as well as by HTS analyses. MAIN CONCLUSIONS Wide virus diversity was found by different methodologies and high frequency of rhinovirus occurrence was confirmed in population in winter, showing its relevance for public health.

Tracking viral particles in the intestinal contents of the American bullfrog, Lithobates catesbeianus, by Transmission Electron Microscopy / Pesquisa de partículas virais em conteúdo intestinal de rãs-touro americanas, Lithobates catesbeianus, por meio da microscopia eletrônica de transmissão

Feces are an important viral agent elimination route for infected carrier animals and in aquatic organisms these pathogenic agents can very rapidly propagate due to the habitation environment. The objective of this work is to track viral particles in the intestinal contents of bullfrogs (Lithobates catesbeianus) from five commercial frog farms in the region of Vale do Paraíba, in the State of São Paulo, Brazil, using negative contrast transmission electron microscopy (TEM). The Coronaviridae, Paramyxoviridae, Parvoviridae and Herpesviridae families were observed and photographed in specimens. This work emphasizes the importance of adopting sanitary measures in commercial farms and confirms that observing feces by TEM is an efficient and rapid diagnostic tool for detecting viral agents.(AU)

Sabendo-se que as fezes são uma importante via de eliminação de agentes virais pelos animais portadores e que, por estarem na água, os agentes patogênicos podem se propagar mais rapidamente, objetivou-se a pesquisa de vírus em conteúdo intestinal de rãs-touro (Lithobates catesbeianus) de cinco ranários comerciais na região do Vale do Paraíba, no estado de São Paulo, pela técnica de microscopia eletrônica de transmissão. As famílias Coronaviridae, Paramixoviridae, Parvoviridae e Herpesviridae foram observadas e fotografadas. Este trabalho ressalta a importância da adoção de medidas sanitárias nas criações, além da confirmação de que a observação de fezes pela microscopia eletrônica de transmissão é uma eficiente ferramenta de diagnóstico rápido para agentes virais.(AU)

Tracking viral particles in the intestinal contents of the American bullfrog, Lithobates catesbeianus, by Transmission Electron Microscopy / Pesquisa de partículas virais em conteúdo intestinal de rãs-touro americanas, Lithobates catesbeianus, por meio da microscopia eletrônica de transmissão

Feces are an important viral agent elimination route for infected carrier animals and in aquatic organisms these pathogenic agents can very rapidly propagate due to the habitation environment. The objective of this work is to track viral particles in the intestinal contents of bullfrogs (Lithobates catesbeianus) from five commercial frog farms in the region of Vale do Paraíba, in the State of São Paulo, Brazil, using negative contrast transmission electron microscopy (TEM). The Coronaviridae, Paramyxoviridae, Parvoviridae and Herpesviridae families were observed and photographed in specimens. This work emphasizes the importance of adopting sanitary measures in commercial farms and confirms that observing feces by TEM is an efficient and rapid diagnostic tool for detecting viral agents...

Sabendo-se que as fezes são uma importante via de eliminação de agentes virais pelos animais portadores e que, por estarem na água, os agentes patogênicos podem se propagar mais rapidamente, objetivou-se a pesquisa de vírus em conteúdo intestinal de rãs-touro (Lithobates catesbeianus) de cinco ranários comerciais na região do Vale do Paraíba, no estado de São Paulo, pela técnica de microscopia eletrônica de transmissão. As famílias Coronaviridae, Paramixoviridae, Parvoviridae e Herpesviridae foram observadas e fotografadas. Este trabalho ressalta a importância da adoção de medidas sanitárias nas criações, além da confirmação de que a observação de fezes pela microscopia eletrônica de transmissão é uma eficiente ferramenta de diagnóstico rápido para agentes virais...

Characterization of parvoviruses from domestic cats in Brazil.

To characterize Feline parvovirus (FPV) circulating in domestic cats in Brazil, 51 fecal samples from unvaccinated domestic cats were collected during 2004-2005. Six parvoviruses were characterized by hemagglutination (HA) assay at different pH values and temperatures and by polymerase chain reaction (PCR) using different pairs of primers. However, data obtained from HA and PCR did not allow the discrimination between FPV and Canine parvovirus (CPV). Two regions of the VP2 capsid gene (1,171-bp fragment) involved in controlling canine and feline host range were sequenced; 9 synonymous and 10 non-synonymous nucleotide substitutions were detected. All samples were confirmed as FPV by nucleotide sequencing, but 3 feline samples had amino acid changes at residues 93, 375, and 426, which are present in canine strains. The phylogenetic tree built based on nucleotide sequences showed that Brazilian feline samples form a cluster distinct from other parvoviruses deposited in GenBank. Taken together, the findings reinforce the importance of monitoring the continuous evolution of CPV and FPV in the feline population in Brazil.

Sequence diversity of TT virus in geographically dispersed human populations.

TT virus (TTV) is a newly discovered DNA virus originally classified as a member of the Parvoviridae. TTV is transmitted by blood transfusion where it has been reported to be associated with mild post-transfusion hepatitis. TTV can cause persistent infection, and is widely distributed geographically; we recently reported extremely high prevalences of viraemia in individuals living in tropical countries (e.g. 74% in Papua New Guinea, 83% in Gambia; Prescott & Simmonds, New England Journal of Medicine 339, 776, 1998). In the current study we have compared nucleotide sequences from the N22 region of TTV (222 bases) detected in eight widely dispersed human populations. Some variants of TTV, previously classified as genotypes 1a, 1b and 2, were widely distributed throughout the world, while others, such as a novel subtype of type 1 in Papua New Guinea, were confined to a single geographical area. Five of the 122 sequences obtained in this study (from Gambia, Nigeria, Papua New Guinea, Brazil and Ecuador) could not be classified as types 1, 2 or 3, with the variant from Brazil displaying only 46-50% nucleotide (32-35% amino acid) sequence similarity to other variants. This study provides an indication of the extreme sequence diversity of TTV, a characteristic which is untypical of parvoviruses.

Human parvovirus B 19 infection and hydrops fetalis in Rio de Janeiro, Brazil

Formalin-fixed paraffin embedded lung and liver tissue from 23 cases of non immune hydrops fetalis and five control cases, in which hydrops were due to syphilis (3) and genetics causes (2), were examined for the presence of human parvovirus B19 by DNA hybridisation. Using in situ hybridisation with a biotynilated probe one positive case was detected. Using 32 P-labelled probes in a dot blot assay format, five further positives were obtained. These were all confirmed as positive by a nested polymerase chain reaction assay. Electron microscopy revealed virus in all these five positive cases. The six B19 DNA positive cases of hydrops fetalis were from 1974, 1980, 1987 and 1988, four of which occurred during the second half of the year, confirming the seasonality of the disease.

Outbreaks of porcine parvovirus disease in Panama.

The first recorded isolation of porcine parvovirus (PPV) in Panama is described. The outbreaks of PPV disease were characterised by a high prevalence of mummified foetuses, stillborn and weak pigs and a common source of exposure. Diagnosis was based on virus isolation and by demonstrating viral antigen in lungs of affected foetuses. Six farms in four different provinces were involved. Rapid control of the epizootic was achieved through the use of an inactivated PPV vaccine in the affected farms.

Viral haemorrhagic disease of rabbits in Mexico: epidemiology and viral characterization.

A fatal disease of rabbits was first reported in the People's Republic of China in 1984. Since 1986, the disease has been reported in most countries of Europe and in the Republic of Korea. In 1989 a similar disease, presumably linked to the importation of rabbit meat from the People's Republic of China, spread rapidly through ten states in Mexico; it was eradicated during the same year by "stamping-out" measures. In Mexico, as was the case in other outbreaks, morbidity and mortality reached 80-90% with few clinical signs. In pathogenesis studies, the primary sites of replication were in the small intestinal crypt and villous epithelium, hepatocytes and splenic lymphocytes. Many organs, including the lung and kidney, contained acutely infarcted tissue and haemorrhages resulting from a terminal disseminated intravascular coagulopathy. The disease and the characteristics of the virus isolated in Mexico are similar to isolates from Europe and the Republic of Korea. The comparative morphologic, from Europe and the Republic of Korea. The comparative morphologic, immunologic, and in situ nucleic acid hybridization evidence for a parvovirus aetiology are summarized.

Isolamento de parvovirus canino no Brasil / Isolation of canine parvovirus in Brazil

Foram examinadas 56 amostras de fezes, 10 de raspado de mucosa intestinal de 2 amostras de material vomitado de cäes doentes, além de 7 amostras de fezes de cäes normais, com o objetivo de isolar-se o parvovirus canino durante a epizootia de gastoenterite hemorrágica ocorrida em Säo Paulo. Das 56 amostras de fezes de cäes doentes foram isolados virus de 24 (42,8%) e das 7 amostras de fezes de cäes clinicamente normais foi isolado virus de apenas 1 (14,3%). Tanto o material vomitado quanto do raspado de mucosa intestinal obteve-se o isolamenteo de parvovirus de uma amostra apenas, correspondendo a 50% e 10%, respectivametne. A reaçäo de hemaglutinaçäo, realizada com os váios tipos de materiais clínicos, demonstrou concordância de 96,3% com o isolamento de virus nas 27 amostras positivas, evidenciando a grande utilidade prática desta reaçäo como método de triagem para determinar, nos casos de diarréia em cäes, a presença de particulas virais com esta caracteristica hemaglutinantes. Em 15 amostras de suspensäo fecal e em uma das culturas positivas foi observada, a microscopia eletrônica, a presença de particulas virais com a s características morfológicas dos parvovirus isolados e o parvovirus felino foi demonstrada pela reaçäo de inibiçäo de hemaglutinaçäo e pela imunofluorescência em 6 das amostras isoladas