DIAGNÓSTICO MOLECULAR DA RAIVA A PARTIR DE DIFERENTES SEGMENTOS DO SISTEMA NERVOSO CENTRAL DE EQUINOS DO MATO GROSSO DO SUL

Abstract

Rabies is an anthropozoonosis caused by Lyssavirus rabies, leading to acute, progressive, and fatal encephalitis in nearly all cases, constituting a global public health concern. The gold standard laboratory method for rabies diagnosis is direct immunofluorescence (IFD) combined with the biological test (PB) through intracerebral inoculation in mice, which has been increasingly replaced by molecular diagnosis. Reports indicate differences in viral distribution within the central nervous system (SNC) structures of equines, with higher detection rates in the brainstem and spinal cord. Thus, the present study aimed to assess the differences in rabies virus detection across distinct CNS regions, including the cortex, cerebellum, thalamus, hippocampus, and cervical, thoracic, and lumbar spinal cord segments, using real-time reverse transcription polymerase chain reaction (RT-qPCR) to quantify the nucleic acid present in each SNC segment of equines. These findings were analyzed alongside the DIF results obtained separately from each SNC segment. Between 2022 and 2024, a total of 33 SNC samples were collected from equines exhibiting neurological symptoms, with varying disease onset and fatality (euthanasia or natural death), and suspected of rabies. These samples were submitted to IAGRO-MS for diagnostic evaluation. Upon receipt, rabies virus RNA was immediately extracted using Trizol to prevent material degradation due to repeated freeze-thaw cycles. All samples were subsequently analyzed by RT-qPCR. Among the 33 samples collected, only twelve (12) tested positive for rabies. Of these, only one (1) sample contained all seven SNC fragments required for diagnosis. In this sample, the lumbar spinal cord segment exhibited the lowest Cq value (25.19), indicating a difference of at least 3.8 Cq compared to the other SNC fragments. This finding suggests that the lumbar spinal cord portion in this sample harbored an eightfold higher viral load than the other SNC segments. In samples two (2) and six (6), the cervical spinal cord exhibited the lowest Cq value, whereas in samples five (5), seven (7), nine (9), ten (10), and twelve (12), the cortex had the lowest Cq. No consistent pattern of viral load distribution was observed in the equine SNC. Some samples showed higher viral loads in the lumbar spinal cord, while others had greater concentrations in the cortex, thalamus, or cerebellum. These findings reinforce the importance of collecting multiple SNC fragments to ensure accurate rabies diagnosis.