Reliable detection of St. Louis encephalitis virus by RT-nested PCR.
BACKGROUND
St. Louis encephalitis virus (SLEV) is a re-emerging arbovirus in South America, with reported cases in humans in Argentina and Brazil. This fact indicates that there is an urgent need to increase the current knowledge about this virus in order to control and prevent future cases. Exhaustive epidemiological and laboratory investigation is required to ensure fast, accurate identification of the viral agent and allow prompt surveillance action by health authorities. Herein, we report the development of a species-specific RT-nested PCR to detect SLEV.


METHODS
After selecting the SLEV genomic region providing the greatest information on the natural genetic variability of this virus, degenerated oligonucleotide primers were designed to amplify a 234-bp fragment of the envelope gene from nine SLEV strains (Parton, BeH356964, SPAN11916, AN9275, AN9124, 78V6507 and 3 SLEV strains obtained from naturally infected mosquito pools).


RESULTS
The method was able to identify the genome of all the SLEV strains tested and did not amplify unrelated RNA viruses, such as yellow fever virus, Ilheus virus, dengue-2 virus, Bussuquara virus, West Nile virus, Japanese encephalitis virus and Murray Valley encephalitis virus. The method was specific and sensitive, with a lower detection limit of < 10 plaque-forming units.


CONCLUSIONS
This molecular assay is a reliable procedure with a wide spectrum for detecting the natural diversity of SLEV and may be useful for ecological studies, clinical and laboratory settings and virological surveillance.
Ré
Viviana
V
Instituto de Virología Dr. J.M. Vanella, Facultad de Ciencias Médicas, Universidad Nacional de Córdoba, Córdoba, Argentina.
Spinsanti
Lorena
L
Farías
Adrián
A
Díaz
Adrián
A
Vázquez
Ana
A
Aguilar
Javier
J
Tenorio
Antonio
A
Contigiani
Marta
M
D000818:Animals; D001118:Argentina / Q000453:epidemiology; D002522:Chlorocebus aethiops; D009033:Culicidae / Q000821:virology*; D004196:Disease Outbreaks; D004665:Encephalitis Virus, St. Louis / Q000235:genetics / Q000302:isolation & purification*; D004674:Encephalitis, St. Louis / Q000453:epidemiology / Q000821:virology; D018067:Flaviviridae / Q000235:genetics; D016679:Genome, Viral; D006801:Humans; D007303:Insect Vectors / Q000821:virology*; D012367:RNA, Viral / Q000032:analysis* / Q000235:genetics; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D014709:Vero Cells / Q000821:virology; D010948:Viral Plaque Assay
D012367:RNA, Viral