Isolation of saint louis encephalitis virus from a horse with neurological disease in Brazil

Abstract

St. Louis encephalitis virus (SLEV) is a causative agent of encephalitis in humans in the Western hemisphere. SLEV is a positive-sense RNA virus that belongs to the Flavivirus genus, which includes West Nile encephalitis virus, Japanese encephalitis virus, Dengue virus and other medically important viruses. Recently, we isolated a SLEV strain from the brain of a horse with neurological signs in the countryside of Minas Gerais, Brazil. The SLEV isolation was confirmed by reverse-transcription RT-PCR and sequencing of the E protein gene. Virus identity was also confirmed by indirect immunofluorescence using commercial antibodies against SLEV. To characterize this newly isolated strain in vivo, serial passages in newborn mice were performed and led to hemorrhagic manifestations associated with recruitment of inflammatory cells into the central nervous system of newborns. In summary this is the first isolation of SLEV from a horse with neurological signs in Brazil.

Figure 1. Analysis of the envelope (E) gene sequence of SLEV.

Phylogenetic tree was constructed using partial E gene sequence by neighbor-joining (NJ) method (using p-distance) of 1,000 bootstrap replicates using Mega 4.0.2. Boostraps values above 60% are shown. The scale bar represents 5% nucleotide sequence divergence. WNV, JEV and DENV-1 were used as the out group.

Figure 2. Survival rate and frequency of clinical signs in newborn mice inoculated with SLEV.

Uninfected control mice (control), first (P1), second (P2), Third (P3), 4^th (P4), 5^th (P5), 6^th (P6), and 7^th (P7) passages are indicated in the X axis.

Figure 3. Newborn mice inoculated with SLEV presenting neurological and circulatory disorders at the 3^rd viral passage.

(A) Bending of the spine (thoracic kyphosis), and limited mobility. (B) Front limb flexed characterizing loss of proprioception and apathy. (C) Necrosis of the distal extremity of the hind limbs and at the tip of the tail. (D) Hyperemia and extensive areas of hemorrhage in the meninges at the rostral part of the brain.

Figure 4. Central nervous system from newborn mice infected with SLEV.

(A) Cerebral cortex from a newborn mouse at the 4^th viral passage with mild hyperemia and mild lympho-histiocytic perivascular cuffs, HE, 600×. (B) Choroid plexus from a newborn mouse at the 4^th viral passage, focally extensive hemorrhage in the ventricular cavity, HE, 200×. (C) Heart from a newborn at the 5^th viral passage, with moderate hyperemia and mild multifocal hemorrhage in the myocardium, HE, 200×. (D) Liver from a newborn at the 6^th viral passage, with severe hyperemia and focal hemorrhage, HE, 200×. (E) Lung from a newborn at the 7^th viral passage, with severe hyperemia, HE, 200×.