Nucleotide sequences of two regions of the genomes of 11 yellow fever virus (YFV) samples isolated from monkeys or humans with symptomatic yellow fever (YF) in Brazil in 2000, 2004, and 2008 were determined with the objective of establishing the genotypes and studying the genetic variation. Results of the Bayesian phylogenetic analysis showed that sequences generated from strains from 2004 and 2008 formed a new subclade within the clade 1 of the South American genotype I. The new subgroup is here designated as 1E. Sequences of YFV strains recovered in 2000 belong to the subclade 1D, which comprises previously characterized YFV strains from Brazil. Molecular dating analyses suggested that the new subclade 1E started diversifying from 1D about 1975 and that the most recent 2004-2008 isolates arose about 1985.

Since 2000, the expansion of Sylvatic Yellow Fever (YF) has been observed in the southeast of Brazil, being detected in areas considered silent for decades. Epizootics in non-human primates (NHPs) are considered sentinel events for the detection of human cases. It is important to report epizootic events that could have impact on the conservation status of susceptible species. We describe the epizootics in NHPs, notified in state of São Paulo, Brazil, between September 2008 to August 2009. Ninety-one epizootic events, involving 147 animals, were reported in 36 counties. Samples were obtained from 65 animals (44.2%). Most of the epizootics (46.6%) were reported between March and April, the same period during which human cases of YF occurred in the state. Biological samples were collected from animals found dead and were sent to Instituto Adolfo Lutz, in São Paulo. Two samples, collected in two counties without an indication for YF vaccination, were positive for the virus. Another 48 animals were associated with YF by clinical-epidemiological linkage with laboratory confirmed cases. Because the disease in human and NHPs occurred in the same period, the detection of the virus in NHPs did not work as sentinel, but aided in the delineation of new areas of risk.

SUMMARYMayaro virus (MAYV) is an arbovirus (Togaviridae: Alphavirus) enzootic in tropical South America and maintained in a sylvan cycle involving wild vertebrates and Haemagogus mosquitoes. MAYV cases occur sporadically in persons with a history of recent activities inside or around forests. This paper reports three cases of MAYV fever detected in men infected in Camapuã, MS, Brazil. Serum samples collected at four days and two months after the onset of the symptoms and examined by hemagglutination inhibition test, revealed monotypic seroconversion to MAYV. Isolation of the virus was obtained from one of the samples by inoculation of the first blood samples into newborn mice. A suspension of the infected mouse brain was inoculated into C6/36 cells culture and the virus was identified by indirect immunofluorescent assay with alphavirus polyclonal antibodies. RT-PCR, performed with RNA extracted from the supernatant of C6/36 infected cells in the presence of alphavirus generic primers as well as specific MAYV primers, confirmed these results. The reported cases illustrate the importance of laboratory confirmation in establishing a correct diagnosis. Clinical symptoms are not always indicative of a disease caused by an arbovirus. Also MAYV causes febrile illness, which may be mistaken for dengue.

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This paper reports the isolation of St. Louis encephalitis virus (SLEV) from a febrile human case suspected to be dengue, in São Pedro, São Paulo State. A MAC-ELISA done on the patient's acute and convalescent sera was inconclusive and hemagglutination inhibition test detected IgG antibody for flaviviruses. An indirect immunofluorescent assay done on the C6/36 cell culture inoculated with the acute serum was positive for flaviviruses but negative when tested with dengue monoclonal antibodies. RNA extracted from the infected cell culture supernatant was amplified by RT-PCR in the presence of NS5 universal flavivirus primers and directly sequenced. Results of BLAST search indicated that this sequence shares 93% nucleotide similarity with the sequence of SLEV (strain-MSI.7), confirmed by RT-PCR performed with SLEV specific primers. Since SLEV was identified as the cause of human disease, it is necessary to improve surveillance in order to achieve early detection of this agent in the state of São Paulo and in Brazil. This finding is also an alert to health professionals about the need for more complete clinical and epidemiological investigations of febrile illnesses as in the reported case. SLEV infections can be unrecognized or confused with other ones caused by an arbovirus, such as dengue.

Rebouças, M. M. CATROXO, M. H. B.; PONGILUPPI, T.; MELO, N. A.; MILANELO, L.; PETRELLA, S.; MARTINS, A. M. C. P. F. & REBOUÇAS, M. M. Identification of poxvirus under transmission electron microscopy during outbreak period in wild birds, in São Paulo, Brazil. Int. J. Morphol., 27(2):577-585, 2009. SUMMARY:Avianpox is a highly contagious disease infecting both commercial and wild birds, causing great damages to breeders and breeding. Caused by DNA viruses of the family Poxviridae, genus Avipoxvirus, if manifest through 3 forms, cutaneous, diphtheric and septicemic. In June 2003, during illegal commercialization of Brazilian birds, 800 wild birds (Paroaria dominicana, Sporophila caerulescens and Sporophila albogularis) were apprehended and being forwarded to the CRAS (Wild Animals Recovery Center), Tietê Ecological Park. After one month, birds presented cutaneous lesions in the beak and feet and anorexia, emaciation, locomotion difficulties, diarrhea, dehydration and death. Among the 800 birds, 500 died and 40 these (15 Paroaria dominicana, 15 Sporophila caerulescens and 10 Sporophila albogularis) were sent to the Electron Microscopy Laboratory of the Biology Institute of São Paulo, SP, to investigate viral agents. Scabs and fragments of skin lesions collected of theses birds were processed for transmission electron microscopy utilizing negative staining (rapid preparation), resin embedding and immunocitochemistry techniques. Under the transmission electron microscopy in all the analyzed samples it was visualized two types of poxvirus particles, M form, with regular spaced thread-like ridges comprising the exposed surface, measuring 280 x 230 nm; C form or stain-penetrated particle showing the dumbbell-shaped core surrounded by the outer envelope, measuring 360 x 330 nm. In the ultrathin sections obtained, three types of intracytoplasmic inclusion bodies were encountered: type A or Bollinger body, outlined by membrane, containing in its interior a great number of mature particles, measuring 200 x 300 nm, revealing the inner dumbbell-shaped core, two lateral bodies and an external envelope. In the type B electron dense inclusions bodies, viral particles budding of dense amorphous material were observed. Fibrillar inclusions constituted by groups of fibrils or lamellae were disposed in groups witch vary from 2 up to 5 e sometimes showed cross striations. A great number of vesicles, on the average measuring 1000 x 650 nm, containing in its interior granular material were also visualized. The nuclei were deformed and showed a marginalized chromatin. In the immunocytochemistry technique, the antigen-antibody was strongly enhanced by the dense gold particles over the viruses. CATROXO, M. H. B.; PONGILUPPI, T.; MELO, N. A.; MILANELO, L.; PETRELLA, S.; MARTINS, A. M. C. P. F. & REBOUÇAS, M. M. Identification of poxvirus under transmission electron microscopy during outbreak period in wild birds, in São Paulo, Brazil. Int. J. Morphol., 27(2):577-585, 2009.

SUMMARYAfter detecting the death of Howlers monkeys (genus Alouatta) and isolation of yellow fever virus (YFV) in Buri county, São Paulo, Brazil, an entomological research study in the field was started. A YFV strain was isolated from newborn Swiss mice and cultured cells of Aedes albopictus -C6/36, from a pool of six Haemagogus (Conopostegus) leucocelaenus (Hg. leucocelaenus) mosquitoes (Dyar & Shannon) collected at the study site. Virus RNA fragment was amplified by RT-PCR and sequenced. The MCC Tree generated showed that the isolated strain is related to the South American I genotype, in a monophyletic clade containing isolates from recent 2008-2010 epidemics and epizootics in Brazil. Statistical analysis commonly used were calculated to characterize the sample in relation to diversity and dominance and indicated a pattern of dominance of one or a few species. Hg. leucocelaenus was found infected in Rio Grande do Sul State as well. In São Paulo State, this is the first detection of YFV in Hg. leucocelaenus.

SUMMARY:Papillomaviruses, belonging to the Papillomaviridae family, are small oncogenic viruses, causing papillomas and fibropapillomas in the mucosal and cutaneous epithelia of several animals. In bovine species, thirteen types (BPV 1-13) were characterized to date. In this study, the occurrence of papillomatosis in four outbreaks in cattle herds, coming from Brazilian states were registered. The papillomatous lesions were found located in the teats, udders, head and neck. Under the transmission electron microscope, by the negative staining technique, it was possible to visualized rounded-format papillomavirus, with icosahedral symmetry, characterized as "full" and "empty" particles, measuring on average 60 nm in diameter, in all the 40 samples observed of skin lesion fragments. The ultrathin sections revealed the presence of groups of viral, intranuclear, rounded particles measuring 35 nm in diameter and tubular particles with a diameter of 35-39 nm. At immunoelectron microscopy technique, positivity obtained was marked by the presence of aggregates of viral particles formed by the antigen-antibody interaction. In the immunocytochemistry technique, the antigen-antibody reaction showed colloidal gold particles evenly distributed over the surface of the virus. These results showed the importance of the transmission electron microscopy techniques in the diagnosis of bovine papillomatosis that can be used in routine procedures to identify viral agent of this important disease.