Discrimination between dog-related and vampire bat-related rabies viruses in Brazil by strain-specific reverse transcriptase-polymerase chain reaction and restriction fragment length polymorphism analysis.
BACKGROUND
There is a geographical overlap between the two main rabies epidemiological cycles maintained by dogs and vampire bats in Latin America. The geographical and temporal coincidence of rabies outbreaks of respective origins is not unusual in rural areas of Latin America. These circumstances make it difficult to discriminate the intraspecies and interspecies transmission pathways of rabies.


OBJECTIVE
This study was conducted to develop techniques to discriminate dog-related and vampire bat-related rabies virus isolates (DRRV and VRRV, respectively) in Brazil.


METHODS
The 1396 nucleotides of the nucleoprotein gene of a total of 27 DRRV and VRRV were sequenced. Strain-specific (SS) primers were developed based on these sequences. Forty-nine rabies virus strains isolated from animals and humans in several parts of Brazil were examined by reverse transcriptase-polymerase chain reaction (RT-PCR) with SS primers. These rabies viruses were also amplified by RT-PCR with general rabies primers and the PCR products were cut by three restriction enzymes, Blp I, Bsu36 I and BspE I.


RESULTS
All the DRRV and VRRV were distinguished by RT-PCR with SS primers. The PCR products obtained from DRRV were cut at one site by Blp I, but not by Bsu36 I. The PCR products obtained from VRRV were cut at one or two sites by Bsu36 I, but not by Blp I. Blp I and Bsu36 I clearly discriminated DRRV and VRRV in restriction fragment length polymorphysim (RFLP) assays. The results of SS RT-PCR and RFLP were consistent.


CONCLUSIONS
SS RT-PCR and RFLP assays have been developed for determining the origins of rabies virus isolates in Brazil. These assays are simple and rapid, and will be useful for identifying the rabies virus reservoirs of field isolates in Brazil, especially when used together.
Ito
Mikako
M
Department of Preventive Veterinary Medicine and Animal Health, Nihon University School of Veterinary Medicine, 1866 Kameino, Fujisawa, 252-8510, Japan.
Itou
Takuya
T
Shoji
Youko
Y
Sakai
Takeo
T
Ito
Fumio H
FH
Arai
Yohko T
YT
Takasaki
Tomohiko
T
Kurane
Ichiro
I
D000595:Amino Acid Sequence; D000818:Animals; D001938:Brazil / Q000453:epidemiology; D002371:Cat Diseases / Q000821:virology; D002415:Cats; D002417:Cattle; D002418:Cattle Diseases / Q000821:virology; D002685:Chiroptera / Q000821:virology*; D017931:DNA Primers; D015252:Deoxyribonucleases, Type II Site-Specific; D004196:Disease Outbreaks; D004283:Dog Diseases / Q000453:epidemiology / Q000821:virology*; D004285:Dogs; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D010802:Phylogeny; D012150:Polymorphism, Restriction Fragment Length; D011818:Rabies / Q000453:epidemiology / Q000635:transmission / Q000662:veterinary* / Q000821:virology*; D011820:Rabies virus / Q000145:classification* / Q000302:isolation & purification; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012689:Sequence Homology, Nucleic Acid; D012756:Sheep; D012757:Sheep Diseases / Q000821:virology; D013045:Species Specificity; D013552:Swine; D013553:Swine Diseases / Q000821:virology
D017931:DNA Primers; D015252:Deoxyribonucleases, Type II Site-Specific; C053919:TCCGGA-specific type II deoxyribonucleases; C062194:endodeoxyribonuclease SauI