Stacey Human

Lineage 2 West Nile Virus as Cause of Fatal Neurologic Disease in Horses, South Africa

Lineage 2 WNV may be missed as a cause of neurologic infections in horses and humans in this region.

A highly sensitive method for the detection and genotyping of West Nile virus by real-time PCR.

In recent years, West Nile virus has been responsible for outbreaks in regions where it has not previously been found. Five genetic lineages with specific geographic distributions exist. Recent outbreaks of WNV associated with the introduction of lineage 1 strains into the western hemisphere, together with the emergence of lineage 2 WNV in Central Europe, has highlighted the potential for spread of pathogenic WNV strains beyond their expected geographical boundaries. Therefore, genotyping of WNV strains may have important applications in surveillance and epidemiology. We report here the development of a nested real-time PCR for the detection and genotyping of WNV strains by means of dissociation-curve analysis, using fluorescence resonance energy transfer (FRET) probe technology. Eight WNV strains, representing three lineages were tested and correctly genotyped at a detection limit of 0.07 viral genome copies/ml in one-step real-time RT-PCR or 7x10(-16) viral genome copies/ml in a nested real-time PCR. WNV could be identified and typed in serum and brain specimens from a human and horse with neurological disease. To our knowledge, this is the first assay designed for the simultaneous detection and genotyping of WNV by rapid, sensitive real-time PCR which may be implemented in diagnostic and epidemiology laboratories.

Fatal neurologic disease and abortion in mare infected with lineage 1 West Nile virus, South Africa.

In 2010, lineage 1 West Nile virus was detected in South Africa in the brain of a pregnant mare that succumbed to neurologic disease and in her aborted fetus, suggesting an association with abortion in horses. All West Nile virus strains previously detected in horses and humans in South Africa were lineage 2.

Transmission of West Nile virus during horse autopsy.

To the Editor: West Nile virus (WNV) circulates mainly in birds and ornithophilic mosquitoes. Humans and horses are considered incidental, dead-end hosts (1). Fever, rash, arthralgia, and myalgia develop in ≈20% of cases in humans; severe neurologic disease may develop in <1% (1). In horses, 20% of infections result in clinical disease, of which ≈90% involve neurologic disease with ataxia, weakness, recumbency, muscle fasciculation, and high death rates (30%) (2).

Sindbis and Middelburg Old World Alphaviruses Associated with Neurologic Disease in Horses, South Africa.

Old World alphaviruses were identified in 52 of 623 horses with febrile or neurologic disease in South Africa. Five of 8 Sindbis virus infections were mild; 2 of 3 fatal cases involved co-infections. Of 44 Middelburg virus infections, 28 caused neurologic disease; 12 were fatal. Middelburg virus likely has zoonotic potential.

Full-Genome Sequence of a Neuroinvasive West Nile Virus Lineage 2 Strain from a Fatal Horse Infection in South Africa

ABSTRACT We report here the complete genome sequence of a lineage 2 West Nile virus (WNV) strain that resulted in fatal neurological disease in a horse in South Africa. Several recent reports exist of neurological disease associated with lineage 2 WNV in humans and horses in South Africa and Europe; however, there are a lack of sequencing data from recent fatal cases in Southern Africa, where these strains likely originate. A better understanding of the genetic composition of highly neuroinvasive lineage 2 strains may facilitate the identification of putative genetic factors associated with increased virulence.

Pathology of fatal lineage 1 and 2 West Nile virus infections in horses in South Africa

Since 2007, West Nile virus (WNV) has been reported in South African horses, causing severe neurological signs. All cases were of lineage 2, except for one case that clustered with lineage 1 viruses. In the present study, gross and microscopic lesions of six South African lineage 2-infected horses and the one lineage 1 case are described. Diagnoses were confirmed by real-time reverse-transcriptase polymerase chain reaction (RT-PCR) of central nervous system (CNS) tissue and one by RT-PCR of a brain virus isolate. The CNS of all cases was negative by RT-PCR or immunohistochemistry (IHC) for African horse sickness (AHS), equine encephalosis virus, equine herpes viruses 1 and 4, other zoonotic flaviviruses, alphaviruses, and shunivirus, and either by immunofluorescence or IHC for rabies. Gross visceral lesions were nonspecific but often mimicked those of AHS. The CNS histopathology of WNV lineage 2 cases resembled the nonsuppurative polioencephalomyelitis reported in the Northern Hemisphere lineage 1 and recent Hungarian lineage 2 cases. Occasional meningitis, focal spinal ventral horn poliomalacia, dorsal and lateral horn poliomyelitis, leucomyelitis, asymmetrical ventral motor spinal neuritis and frequent olfactory region involvement were also seen. Lineage 2 cases displayed marked variations in CNS lesion severity, type and distribution, and suggested various viral entry routes into the CNS, based on findings in experimental mice and hamsters. Lineage 1 lesions were comparable to the milder lineage 2 cases. West Nile virus IHC on CNS sections with marked lesions from all cases elicited only two antigen-positive cells in the olfactory cortex of one case. The presence in the CNS of T-lymphocytes, B-lymphocytes, plasma cells and macrophage-monocytes was confirmed by cluster of differentiation (CD) 3, CD20, multiple myeloma oncogene 1 (MUM1) and macrophage (MAC) 387 IHC.