C56bl 6

Animal model
Eight-week-old male C56BL/6 (Jackson laboratory # 000664), (wild type, weighing approximately 20–25 grams were used. The animals were obtained from The Jackson Laboratory (Maine, United States of America), and maintained by the central vivarium of the University of Mogi das Cruzes (Ceua # 010/2019 / # 006/2017 / 016/2016 and 01/2017). The animals were kept in individual cages, at an average temperature of 22°C ± 1°C, or in a cold chamber, at 4°C ± 1°C, for 7 days (prolonged exposure to cold), with light/dark cycles every 12 hours, in a controlled manner, besides the provision of feed (Nuvilab^®, Nuvital S/A, Colombo, PR) and water ad libitum. UCP1- Cre mice and B6.129(Cg)-Gt(ROSA)26Sortm4(ACTB-tdTomato,-EGFP)Luo/J (mTmG) mice are from Jackson Laboratory. The Ucp1-cre mice were crossed with mTmG mice to remove the flox-flanked resistance cassette. Mice were housed at 23°C in a 12 hr light/dark cycle with free access to normal chow. All experiments used matched littermates. Experimental procedures were approved by the Boston University Institutional Animal Care and Use Committee.

Six to eight-week-old female wild-type C56BL/6 were purchased from Jackson Laboratories (Bar Harbor, ME, USA). All animals were housed in a pathogen-free environment. All virus infections were performed in the ABSL2 or ABSL3 facility in the Galveston National Laboratory (GNL), UTMB. All animal studies were reviewed and approved by the Institutional Animal Care and Use Committee at UTMB and were carried out in accordance with the National Institute of Health guidelines. Animals were anesthetized using an isoflurane precisions variable-bypass vaporizer prior to intranasal virus inoculation with 10³ PFU TC-83 diluted in PBS (N = 10, performed as two separate experiments of N = 5) or PBS as a mock infection (N = 4). Standardized recording of death and disease symptoms was performed using the following definitions: encephalitis, ruffled fur, hunched back, reduced activity, development of discoordination, ataxia, or transient seizures (with the ability to drink and feed); paralysis; and hind limb (hemiplegic) or quadriplegic paralysis (with the inability to reach the feeder or water bottle). Body weight was measured throughout the course of study. Seroconversion was determined by plaque reduction neutralization test (PRNT) at 6 months post infection (m.p.i.).