Subclinical bovine vaccinia: An important risk factor in the epidemiology of this zoonosis in cattle.

Bovine vaccinia (BV) is a zoonosis caused by Vaccinia virus (VACV) that mainly affects lactating cows and dairy farm milkers. The epidemiological role(s) of other cattle categories such as dry cows, bulls, and heifers in BV remains unclear. This study was performed to investigate VACV in affected dairy cattle herds and perifocal farms during an outbreak in Brazil. Crusts from lesions of cows' teats were collected from all farms with BV outbreaks. Milk, feces, blood, and serum were collected from symptomatic and asymptomatic lactating cows. Blood and serum were also sampled from other cattle categories (calves, heifers, dry cows, and bulls). The samples were tested for VACV by PCR, and to confirm VACV viability, VACV-positive samples were inoculated in BSC-40 cells and stained using immunoperoxidase. Neutralizing antibodies were investigated using plaque reduction neutralization test. Viral DNA was detected in milk, blood, and feces samples of symptomatic and asymptomatic dairy cows and in blood samples from other cattle categories on farms with and without confirmed BV outbreak. In affected farms, viable virus was identified in feces and milk samples from lactating cows and in blood samples from asymptomatic dry cows. Viable VACV was also identified in feces from lactating cows and one bull's blood sample from perifocal farms. Neutralizing antibodies were detected in 81.6% of the herds affected by BV and in 53.8% of the herds on perifocal farms. The presented data indicate a potential source of viral dissemination, which contributes to the persistence and spread of VACV in the environment.

Bovine vaccinia: Inactivated Vaccinia virus vaccine induces protection in murine model.

Bovine vaccinia (BV), caused by Vaccinia virus (VACV), is a zoonosis characterized by exanthematous lesions on the teats of dairy cows and the milkers' hands. Since 1999, due to the occurrence of many BV outbreaks in dairy farms across all Brazilian regions, there is a need to improve the control and prevention measures of the disease. Vaccination is one of the major tools to prevent viral diseases, and it could be an alternative for BV prevention. The main objective of this study was the development of vaccine formulations against BV using the inactivated VACV strain GP2 as antigen combined with different adjuvants. Potency tests were performed in mice, which were vaccinated with two doses at a 21-day interval, and then challenged with the vaccine homologous virus. VACV strain GP2 inactivated by beta-propiolactone (BPL) in association with adjuvants was effective in inducing a humoral immune response against VACV, as measured by neutralizing antibody (NA) titers, and was variable depending on the adjuvant used in each vaccine formulation. The vaccine formulation containing aluminum hydroxide (AH) associated with saponin as adjuvant induced the production of high NA titers in all vaccinated mice, giving 100% protection in Balb/c murine model after challenge with homologous virus.

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.