Vigilância epidemiológica do vírus do Nilo Ocidental no mundo e no Brasil: relevância da vigilância equina no contexto de saúde única

O vírus do Nilo Ocidental (WNV) é um flavivírus neuropatogênico transmitido por mosquito, mantido na natureza emum ciclo de transmissão zoonótica entre as aves e os mosquitos ornitofílicos, principalmente do gênero Culex. Até a décadade 1990, o WNV era considerado um arbovírus do mundo antigo, mas em 1999 surgiu nos Estados Unidos da Américae se espalhou rapidamente, tornando-se uma grande ameaça à saúde pública. O WNV se adaptou ao ciclo envolvendomosquitos e pássaros americanos e chegou à América Central e do Sul nos anos subsequentes. Em 2003, o Sistema Nacionalde Vigilância da Febre do Nilo Ocidental no Brasil foi criado com base na triagem sorológica de animais sentinelas evetores, conforme recomendado pela Organização Pan-Americana da Saúde (OPAS) e pela Organização Mundial daSaúde (OMS). Desde 2008, evidências sorológicas de infecção por WNV em equinos brasileiros têm sido relatadas e acirculação do WNV monitorada por meio de triagem sorológica de cavalos sentinelas, além da notificação de casos deencefalomielite. Os equinos são altamente suscetíveis ao WNV e surtos de doenças neurológicas geralmente precedemcasos humanos. Nesse sentido, a vigilância equina tem sido essencial para fornecer um alerta precoce às autoridades desaúde pública e animal em vários países, incluindo o Brasil. Isso demonstra a necessidade de programas de intervençãoem saúde pública e animal para alocar recursos e conscientizar os médicos veterinários sobre seu papel em processos devigilância humana que envolvam equinos. Nesta revisão, é discutida a importância da vigilância equina e dos médicosveterinários como linha de frente na vigilância humana no Brasil e no mundo, no contexto de saúde única.West Nile virus (WNV) is a neurovirulent mosquito-borne Flavivirus that is maintained in nature by a zoonotic transmissioncycle between avian hosts and ornithophilic mosquito vectors, mostly from the Culex genus. Until the 1990s, WNV wasconsidered to be an old-world arbovirus, but in 1999, WNV emerged in the United States (US) and spread rapidly, becoming amajor threat to public health. WNV adapted to the transmission cycle involving American mosquitoes and birds and reachedCentral and South America in subsequent years. In 2003, the National West Nile Fever Surveillance System was created in Brazilbased on serological screening of animals and sentinel vectors, as recommended by the Pan American Health Organization(PAHO) and the World Health Organization (WHO). Since 2008, serological evidence of WNV infection in Brazilian horseshas been reported, and the circulation of WNV has been monitored through the regular serological screening of sentinel horsesand reporting of encephalomyelitis cases. Horses are highly susceptible to WNV infection, and outbreaks of neurologicaldisease among horses often precede human cases. In this regard, equine surveillance has been essential in providing earlywarning to public and animal health authorities in several countries, including Brazil. This demonstrates the need for animaland public health intervention programs to allocate resources to make veterinarians aware of the role they can play in thehuman surveillance processes by monitoring horses. This review discusses the importance of equine surveillance and the gapthat veterinarians can fill on the front line in human surveillance, in Brazil and worldwide, in the context of “One Health

Vaccinia bovina em vacas secas e lactantes experimentalmente inoculadas com o Vaccina virus

Exportado OPUSMade available in DSpace on 2019-08-13T04:13:04Z (GMT). No. of bitstreams: 1 iza_disserta__o_pronta.pdf: 2795893 bytes, checksum: 411ce477cf274efe925b998f978eeb98 (MD5) Previous issue date: 8A presente dissertação foi dividida em três experimentos cujos objetivos foram reproduzir a vaccínia bovina (VB) em vacas secas e lactantes mestiças através da inoculação experimental com Vaccinia virus amostra GP2 (VACV-GP2). O objetivo do experimento 1 foi estudar a evolução clínico-patológica das lesões e a localização do vírus por imuno-histoquímica em períodos distintos da doença. Nesse experimento, seis vacas secas mestiças, divididas em três grupos, tiveram os tetos escarificados com auxílio de agulha hipodérmica em sítio delimitado, localizado na área central de cada teto. Dois grupos foram eutanasiados e necropsiados em fases diferentes de evolução da doença e em um grupo realizou-se apenas a biopsia dos tetos nos locais onde apareceram lesões. No experimento 2, três vacas mestiças foram inoculadas a fim de analisar o melhor método para escarificação de pele em tetos e a patogenicidade de dois inóculos diferentes contendo VACV-GP2. Uma vaca foi sacrificada no 17º d.p.i., período em que todos os tetos já haviam cicatrizado. Os objetivos do experimento 3 foram realizar os estudos clínico, hematológico e bioquímico associados à infecção e observar os efeitos da imunodepressão artificial e da reinoculação com VACV-GP2 em vacas lactantes previamente infectadas pelo vírus. Esse experimento foi dividido em duas fases, sendo que na fase 1, oito vacas mestiças em fases distintas de lactação foram inoculadas com o VACV e na fase 2, as vacas inoculadas nos experimentos 1 e 3 (parte 1) foram reinoculadas ou imunodeprimidas. Em todos os experimentos os animais foram acompanhados durante 32 dias e foi realizado o exame clínico em dias alternados. Além disso, foram coletadas amostras de sangue, fezes, suabe oral e leite e as técnicas realizadas foram IPMC, soroneutralização e PCR. Nas necropsias, foram coletados diversos tecidos para realização das técnicas de HE e IHQ. No experimento 3 foram acrescentadas as análises de bioquímica sérica e hemograma, além da contagem de células somáticas do leite. De maneira geral, observou-se que o período de incubação do VACV em bovinos é curto, e que o padrão de evolução das lesões ocorreu de maneira similar nos três experimentos, o qual também foi similar ao padrão de evolução de lesões observado em infecções naturais pelo VACV em bovinos. No acompanhamento clínico, foi observado que a infecção experimental do VACV em bovinos causa uma linfoadenopatia local e não foi detectada hipertermia nos bovinos experimentalmente inoculados, nem outras alterações clínicas nesses animais. Alterações histopatológicas significativas foram observadas nos tetos, glândulas mamárias e linfonodos retromamários dos animais inoculados dos experimentos 1 e 2, em três fases diferentes da doença: ulcerativa, crostosa e cicatrização. O VACV foi detectado, através da técnica de IHQ, nos tetos, glândula mamária e linfonodos retromamários dos animais da fase inicial (4º d.p.i) e da fase intermediária (9º d.p.i) da evolução da doença e na fase final, de cicatrização (17º d.p.i). Foi observado também que a mastite pode ser exacerbada pela VB e que houve queda de cerca de 30% na produção de leite nos animais infectados, assim como aumento significativo no número das células somáticas. Em relação ao perfil hematológico, foram observadas linfopenia e neutrofilia, que podem estar associados, respectivamente, com a infecção viral e a inflamação da glândula mamária. Mediante as condições experimentais às quais as vacas foram submetidas, foi possível concluir que a reinfecção pelo VACV pode ocorrer em animais previamente infectados. Suspeita-se também de que o VACV pode persistir e multiplicar em vacas imunodeprimidas, uma vez que houve aumento no título de anticorpos nesse grupo de animais. Por fim, alguns resultados do presente estudo sugerem que o VACV pode disseminar-se sistemicamente no organismo do bovino, uma vez que o DNA viral foi detectado nas lesões de mucosa oral.This dissertation was divided into three experiments whose aims were to reproduce the bovine vaccínia (BV) in crossbred dairy and dry cows through the experimental inoculation with Vaccinia virus Guarani P2 (VACV-GP2). Experiment 1 studied the clinical and pathological evolution of the lesions and the localization of the virus by immuno-histochemistry in different stages of the disease. In this experiment, the teats of six crossbred dry cows, divided into three groups, were scarified with a hypodermic needle in the central area of each teat. Two groups were euthanized and necropsied in different stages of the disease, while in a third group, a biopsy was performed in the inoculated area of the teats In the experiment 2, three crossbred cows were inoculated in order to analyze different scarification methods and the pathogenicity of two different inocula with VACV-GP2. One cow was euthanized on the 17th day post-inoculation (d.p.i), period in which the lesions were healed. The aims of experiment 3 were to carry out clinical, hematological and biochemical analysis associated to the infection and to observe the effects of immunodepression and re-infection with VACV-GP2 in dairy cows previously infected by VACV. The experiment was divided into two parts. In part 1, eight crossbred dairy cows were inoculated with VACV-GP2 and in part 2, the cows from experiment 1 and 3 (part 1) were immunodepressed or reinfected. All animals in experiments 1, 2 and 3 were observed for 32 days and clinical examination was done every other day. Moreover, blood, faeces, oral swab and milk samples were collected daily and the technics employed were IPMA, seroneutralization and PCR. Several tissues were collected in necropsies and analysed through HE and IHC. Biochemical and hematological analysis and somatic cell count (SCC) in milk were done only in experiment 3. In all experiments it was observed that the incubation period was short and the evolution of the lesions was similar among the animals experimentally infected. Moreover, these lesions were also similar to the ones in cows naturally infected. Through clinical monitoring, it was observed that experimental infeccion of VACV in bovines causes local lymphadenopathy. Neither hyperthermy nor clinical alterations were detected. In the inoculated animals from experiments 1 and 2, histological alterations were observed in the teats, mammary glands and mammary lymph nodes in the three different phases of the disease, i.e., ulcerative, crust and healing. Through IHC, the VACV was detected in teats, as well as in mammary glands and mammary lymph nodes on the 3th, 9th and 17th d.p.i. During the experiment the mastitis was exacerbated by BV, which was shown by the increase of SCC in milk, and the dairy cows had intense decrease in milk production. lymphopenia and neutrophilia,which may be associated with viral infection and mastitis, respectively, were observed in the hematological analysis. In the experimental conditions which the cows were tested, it was possible to conclude that reinfection by VACV can occur in previously infected animals. There is a suspicion that the VACV can persist and multiply in immunodepressed bovines, once that there was an increase in antibodies titers in these animals. Lastly, some results suggest that the VACV can spread systematically in cattles, since the viral DNA was detected in the oral mucosa lesions

Bovine Vaccinia: Insights into the Disease in Cattle

Bovine vaccinia (BV), caused by Vaccinia virus (VACV), is a zoonosis characterized by exanthematous lesions in the teats of dairy cows and the hands of milkers and is an important public health issue. Severe VACV-induced lesions in the teats and udder of cows and buffaloes could lead to mastitis and other secondary infections, thereby reducing productivity and resulting in economic losses to the dairy industry. In Brazil, BV re-emerged in the late 1990s and is now endemic in most of the Brazilian territory. In the last 15 years, much effort has been made to know more about this disease and its epidemiology, etiologic agents, and interactions with the host and the environment. In this review, we describe the known dynamics of VACV infection in cattle and the viral shedding routes, as well as the relevance of BV for animal and public health

Vaccinia virus Transmission through Experimentally Contaminated Milk Using a Murine Model.

Bovine vaccinia (BV) is a zoonosis caused by Vaccinia virus (VACV), which affects dairy cattle and humans. Previous studies have detected the presence of viable virus particles in bovine milk samples naturally and experimentally contaminated with VACV. However, it is not known whether milk contaminated with VACV could be a route of viral transmission. However, anti-Orthopoxvirus antibodies were detected in humans from BV endemic areas, whom had no contact with affected cows, which suggest that other VACV transmission routes are possible, such as consumption of contaminated milk and dairy products. Therefore, it is important to study the possibility of VACV transmission by contaminated milk. This study aimed to examine VACV transmission, pathogenesis and shedding in mice orally inoculated with experimentally contaminated milk. Thirty mice were orally inoculated with milk containing 107 PFU/ml of VACV, and ten mice were orally inoculated with uncontaminated milk. Clinical examinations were performed for 30 consecutive days, and fecal samples and oral swabs (OSs) were collected every other day. Mice were euthanized on predetermined days, and tissue and blood samples were collected. Nested-PCR, plaque reduction neutralization test (PRNT), viral isolation, histopathology, and immunohistochemistry (IHC) methods were performed on the collected samples. No clinical changes were observed in the animals. Viral DNA was detected in feces, blood, OSs and tissues, at least in one of the times tested. The lungs displayed moderate to severe interstitial lymphohistiocytic infiltrates, and only the heart, tonsils, tongue, and stomach did not show immunostaining at the IHC analysis. Neutralizing antibodies were detected at the 20th and 30th days post infection in 50% of infected mice. The results revealed that VACV contaminated milk could be a route of viral transmission in mice experimentally infected, showing systemic distribution and shedding through feces and oral mucosa, albeit without exhibiting any clinical signs

Histological sections of different tissues from mice experimentally infected with VACV-GP2.

<p><b>Staining using the IHC method (100 μm)</b>. (A) Lung: mild to moderate immunostaining in the lymphocytes cytoplasms (arrows). (B) Spleen: moderate immunostaining in the lymphocytes cytoplasms (arrows). (C) Kidney: mild-to-moderate immunostaining in the cytoplasm of the proximal-convoluted-tubule epithelial cells (arrows). (D) Liver: mild to moderate immunostaining in the cytoplasm of hepatocytes (arrows). (E) Submandibular lymph nodes: mild immunostaining in the lymphocyte cytoplasm (arrows). (F) Ileum: moderate immunostaining in the cytoplasm of Peyer’s patch lymphocytes (arrows) and mild immunostaining in epithelial cells (white arrow).</p

Histological lung sections showing interstitial lymphohistiocytic infiltrate (arrows).

<p>H&E staining. (A)- Mouse lung from the control group (CG) inoculated orally with milk not contaminated with VACV (200 μm). (B)-Mouse lung from the inoculated group (IG) orally inoculated with VACV-contaminated milk (200 μm).</p

Susceptibility of Vaccinia Virus to Chemical Disinfectants

Vaccinia virus (VACV) is the cause of bovine vaccinia (BV), an emerging zoonotic disease that affects dairy cows and milkers. Some chemical disinfectants have been used on farms affected by BV to disinfect cow teats and milkers' hands. To date, there is no information about the efficacy of disinfectants against VACV. Therefore, this study aimed to assess the virucidal activity of some active disinfectants commonly used in the field. Sodium hypochlorite, quaternary ammonium combined with chlorhexidine, and quaternary ammonium combined with glutaraldehyde were effective in inactivating the virus at all concentrations tested. Iodine and quaternary ammonium as the only active component were partially effective. The presence of bovine feces as organic matter and light decreased the effectiveness of sodium hypochlorite. These results show that an appropriated disinfection and asepsis of teats and hands may be helpful in the control and prevention of BV and other infections with VACV