Iara A. Borges

Vaccinia Virus Infection in Monkeys, Brazilian Amazon

To detect orthopoxvirus in the Brazilian Amazon, we conducted a serosurvey of 344 wild animals. Neutralizing antibodies against orthopoxvirus were detected by plaque-reduction neutralizing tests in 84 serum samples. Amplicons from 6 monkey samples were sequenced. These amplicons identified vaccinia virus genetically similar to strains from bovine vaccinia outbreaks in Brazil.

Vaccinia Virus Zoonotic Infection, São Paulo State, Brazil

To the Editor: Since 1999, vaccinia virus (VACV) has been isolated frequently from dairy cattle and humans (1–3). During bovine vaccinia outbreaks, VACV can be transmitted to farmers and those who milk cows; it frequently causes lesions on the hands and forearms. Bovine vaccinia causes economic losses and affects public health services in Brazil (1–4). One of the first VACV viruses isolated during Brazilian bovine vaccinia outbreaks was Aracatuba virus (ARAV), which was collected in Sao Paulo State, and since that time, other VACVs have been isolated in this state (2,5,6).

Group 2 vaccinia virus, Brazil.

In 2011, vaccinia virus caused an outbreak of bovine vaccinia, affecting dairy cattle and dairy workers in Brazil. Genetic and phenotypic analyses identified this isolate as distinct from others recently identified, thereby reinforcing the hypothesis that different vaccinia virus strains co-circulate in Brazil.

Serologic and Molecular Evidence of Vaccinia Virus Circulation among Small Mammals from Different Biomes, Brazil

Vaccinia virus (VACV) is a zoonotic agent that causes a disease called bovine vaccinia, which is detected mainly in milking cattle and humans in close contact with these animals. Even though many aspects of VACV infection have been described, much is still unknown about its circulation in the environment and its natural hosts/reservoirs. To investigate the presence of Orthopoxvirus antibodies or VACV DNA, we captured small rodents and marsupials in 3 areas of Minas Gerais state, Brazil, and tested their samples in a laboratory. A total of 336 animals were tested; positivity ranged from 18.1% to 25.5% in the 3 studied regions located in different biomes, including the Atlantic Forest and the Cerrado. Analysis of nucleotide sequences indicated co-circulation of VACV groups I and II. Our findings reinforce the possible role played by rodents and marsupials in VACV maintenance and its transmission chain.

Vaccinia virus in household environment during bovine vaccinia outbreak, Brazil.

To the Editor: Several exanthematic vaccinia virus (VACV) outbreaks have affected dairy cattle and rural workers in Brazil and Asia, and have caused economic losses and affected health services (1–3). VACV, the prototype of the genus Orthopoxvirus (OPV), exhibits serologic cross-reactivity with other OPV species and was used during the smallpox eradication campaign (1). Several VACV strains have been isolated during bovine vaccinia outbreaks in Brazil and have been characterized by molecular and biologic methods (3,4). Bovine vaccinia infections in humans are frequently related to occupational contact with sick animals during milking but have never been shown to be associated with fomites or indoor environments (1,3). In August 2011, a bovine vaccinia outbreak was reported in Carangola County, Minas Gerais State, Brazil. During this outbreak, several farms were affected, and the outbreak involved humans and dairy cattle. A 41-year-old man (patient 1) who worked on a farm (20°36′30.7′′S, 42°17′53.9′′W) was hospitalized. He had painful lesions on the hands, high fever, lymphadenopathy, malaise, and fainting episodes. This patient reported recent contact with sick animals on the farm during milking. At the same time, a 57-year-old man (patient 2), the owner of the farm, had a lesion on the right hand. This infection was also related to occupational exposure. Some days after the appearance of the hand lesion, this patient presumably inoculated himself at the site of an abrasion he had recently received on his nose. This resulted in development of a large and painful lesion. This patient reported milking cows daily. He had been vaccinated against smallpox before 1977. A total of 5 humans and 15 cows were involved in this outbreak on 5 farms. Clinical samples were obtained from the 2 patients and from 3 sick cows. Dried swab specimens from lesions were soaked in 200 μL of phosphate-buffered saline containing penicillin (200 U/mL), amphotericin B (4 μg/mL), and gentamicin (100 μg/mL); homogenized, and centrifuged at 2,000 × g for 3 min. The supernatants were used for molecular diagnosis and virus isolation (3,5,6). Supernatants were tested by using OPV-specific PCRs that targeted the C11R gene, which encodes viral growth factor (vgf), and the A26L gene, which encodes A-type inclusion (ATI) protein. Samples from the 2 patients were positive for vgf and ATI (7). At least 1 sample (blood or scabs) from each sick animal was also positive by PCR. To assess the risk for virus spread in indoor environments, we collected swab specimens from several objects, including doorknobs, bathroom surfaces, and the pillow of patient 2. The pillow was positive for vgf and ATI by PCR. To isolate the virus, we infected monolayers of BSC-40 cells cultured in a 6-well plate with sample supernatants and incubated the cells at 37°C for 72 h or until a cytopathic effect was detected (3,5,6). We isolated virus from a sample from patient 1 and from an environmental sample (the pillow of patient 2), which showed positive results in the molecular diagnostic assays. To confirm that the isolated VACV was the OPV involved in this outbreak, we sequenced partial fragments of the A56R and A26L genes from the isolated virus. Fragments obtained were directly sequenced in both orientations in triplicate (MegaBACE 1000 Sequencer; GE Healthcare, Little Chalfont, UK). Sequences were aligned with published OPV sequences in GenBank by using the ClustalW (www.clustal.org/) method and manually aligned by using MEGA version 4.0 (Arizona State University, Phoenix, AZ, USA). VACV molecular signatures of 18-nt and 12-nt deletions were observed in the A56R and A26L genes, respectively. Phylogenetic trees (Figure), which were constructed by using the neighbor-joining method, the Tamura-Nei model of nucleotide substitutions, and 1,000 bootstrap replicates in MEGA 4.0, demonstrated that this isolate clustered with other group 1 VACV isolates from Brazil. We named this isolate Carangola virus. Figure Phylogenetic trees based on orthopoxvirus nucleotide sequences, including vaccinia virus (VACV) from Brazil (VACV-BR). Phylogenetic analysis was performed for A56R (A) and A26L (B) gene sequences and grouped VACV-BR strains into 2 branches: group 1 (red ... We isolated VACV from an indoor environment during a bovine vaccinia outbreak. VACV infections have been frequently associated with occupational activities, primarily direct contact with sick animals (1,3). However, in some cases, the source of the infection is unknown, especially in patients who did not participate in milking activities. Human-to-human transmission has been suggested to have occurred in some bovine vaccinia outbreaks in Brazil, and nosocomial infection has been reported Asia (2,8). Household transmission of VACV has also been described in the United States after contact with lesions of a smallpox vaccinee in the military (9). VACV from Brazil shows long-lasting stability under environmental conditions, especially when associated with organic matter (10). Although the wife of patient 2 did not exhibit any typical clinical symptoms of VAVC infection, we believe that relatives sharing household environments with patients with lesions may be at risk for VACV infection. Isolation of VACV from a household environment raises new questions about nonoccupational risk factors related to bovine vaccinia transmission.

Perfil sorológico, viremia e eliminação do circovírus suíno tipo 2 em animais naturalmente infectados pertencentes a granjas com ou sem a síndrome da refugagem multissistêmica

A virological and serological cross-sectional study (E1) and a longitudinal study (E2) were performed on herds with (G2 and G3) and without (G1) post weaning multisystemic syndrome (PMWS) in Brazil. Blood, serum, nasal and rectal swabs samples were collected of sows, farrowing piglets, nursery, growing and finishing pigs. In E1, were sampled 40 animals in each category (G1a and G2). In E2, (G1b and G3), 35 farrowing piglets were identified and sampled along the production cycle. Porcine circovirus type 2 (PCV2) antibodies were assayed. A PCR was used to detected PCV2 genome in blood and swabs. In E1, sows had high rates of viremic and seropositives animals, with percentage of sows with high antibodies titers greater than G2. Passive antibodies decline occurred between nursery and growing area with increased viral shedding in swabs and subsequent seroconversion in G1. In G2, the passive antibodies decay occurred in nursery, with a reduction in viral shedding. In E2, the decline of maternal immunity occurred between the 1st and 2nd collection in G1b, and between 2nd and 3rd collections in G3. In both herds, the decay of passive immunity coincided with increased viremia and viral shedding; and seroconversion occurred between the 3rd and 4th collection in both herds with decline of viremia. Viremia and viral shedding was detected in all samples days, 42% of animals sampled in E2 were viremic and all tissue samples collected at slaughter were positive for PCV2. This study confirms the persistence of viremia even in the presence of high titers of antibodies and the serological profile in a herd with or without PMWS may be different, especially with regard to the passive immunity duration.

Ubiquitous giants: a plethora of giant viruses found in Brazil and Antarctica.

BackgroundSince the discovery of giant viruses infecting amoebae in 2003, many dogmas of virology have been revised and the search for these viruses has been intensified. Over the last few years, several new groups of these viruses have been discovered in various types of samples and environments.In this work, we describe the isolation of 68 giant viruses of amoeba obtained from environmental samples from Brazil and Antarctica.MethodsIsolated viruses were identified by hemacolor staining, PCR assays and electron microscopy (scanning and/or transmission).ResultsA total of 64 viruses belonging to the Mimiviridae family were isolated (26 from lineage A, 13 from lineage B, 2 from lineage C and 23 from unidentified lineages) from different types of samples, including marine water from Antarctica, thus being the first mimiviruses isolated in this extreme environment to date. Furthermore, a marseillevirus was isolated from sewage samples along with two pandoraviruses and a cedratvirus (the third to be isolated in the world so far).ConclusionsConsidering the different type of samples, we found a higher number of viral groups in sewage samples. Our results reinforce the importance of prospective studies in different environmental samples, therefore improving our comprehension about the circulation anddiversity of these viruses in nature.

Evaluating anti-Orthopoxvirus antibodies in individuals from Brazilian rural areas prior to the bovine vaccinia era

Vaccinia virus naturally circulates in Brazil and is the causative agent of a zoonotic disease known as bovine vaccinia (BV). We retrospectively evaluated two populations from the Amazon and Southeast Regions. BV outbreaks had not been reported in these regions before sample collection. Neutralising antibodies were found in 13 individuals (n = 132) with titres ranging from 100 ≥ 6,400 neutralising units/mL. Univariate analysis identified age and vaccination as statistically significant risk factors in individuals from the Southeast Region. The absence of detectable antibodies in vaccinated individuals raises questions about the protection of smallpox vaccine years after vaccination and reinforces the need for surveillance of Orthopoxvirus in Brazilian populations without evidence of previous outbreaks.

Serological Evidence of Orthopoxvirus Circulation Among Equids, Southeast Brazil

Since 1999 Vaccinia virus (VACV) outbreaks involving bovines and humans have been reported in Brazil; this zoonosis is known as Bovine Vaccinia (BV) and is mainly an occupational disease of milkers. It was only in 2008 (and then again in 2011 and 2014) however, that VACV was found causing natural infections in Brazilian equids. These reports involved only equids, no infected humans or bovines were identified, and the sources of infections remain unknown up to date. The peculiarities of Equine Vaccinia outbreaks (e.g., absence of human infection), the frequently shared environments, and fomites by equids and bovines in Brazilian farms and the remaining gaps in BV epidemiology incited a question over OPV serological status of equids in Brazil. For this report, sera from 621 equids - representing different species, ages, sexes and locations of origin within Minas Gerais State, southeast Brazil – were examined for the presence of anti-Orthopoxvirus (OPV) antibodies. Only 74 of these were sampled during an Equine Vaccinia outbreak, meaning some of these specific animals presented typical lesions of OPV infections. The majority of sera, however, were sampled from animals without typical signs of OPV infection and during the absence of reported Bovine or Equine Vaccinia outbreaks. Results suggest the circulation of VACV among equids of southeast Brazil even prior to the time of the first VACV outbreak in 2008. There is a correlation of OPVs outbreaks among bovines and equids although many gaps remain to our understanding of its nature. The data obtained may even be carefully associated to recent discussion over OPVs history. Moreover, data is available to improve the knowledge and instigate new researches regarding OPVs circulation in Brazil and worldwide.

Silent Orthohantavirus Circulation Among Humans and Small Mammals from Central Minas Gerais, Brazil

New World orthohantaviruses are emerging RNA viruses that cause hantavirus cardiopulmonary syndrome (HCPS). These viruses are a burden to public health around the world with a lethality rate of around 60%. In South America, rodents of Sigmodontinae subfamily are the main reservoirs of orthohantaviruses. We described a serosurvey for orthohantaviruses circulation in an apparently healthy human population and small mammals from rural areas in Central Minas Gerais State, Brazil. A total of 240 individuals and 50 small mammals (26 rodents belonging to 10 different species and 24 marsupials from 4 different species) were sampled during 2012–2013. The seroprevalence rates of IgG/IgM antibodies in humans were 7.1 and 1.6%, respectively. Only one rodent, an Oligoryzomys nigripes captured in peridomestic area, tested positive for IgG antibodies and viral RNA. Our findings suggest a silent circulation of orthohantaviruses in a region of intensive agriculture production. The detection of seropositive humans in an area with a lack of previous HCPS reports highlights potential oligosymptomatic cases and the need for surveillance strategies that could reduce the risk of future outbreaks.