[Development of PCR assay for detection of Angiostrongylus cantonensis in Pomacea canaliculata].
OBJECTIVE
To establish a PCR assay for detecting the third-stage larvae of Angiostrongylus cantonensis in Pomacea canaliculata.


METHODS
Polymerase chain reaction primers were designed by the software Lasergene, based on the specific cDNA of the third-stage larvae of A.cantonensis in Genbank. The total RNA was prepared from the third-stage larvae of A.cantonensis and of the snails by TRIzol one-step protocol. Amplification by RT-PCR was carried out following the kit protocol.


RESULTS
RT-PCR assay revealed a clear differentiation between infected and negative snails. When a mixture of the total RNA from the negative snails and the third-stage larvae of A.cantonensis was tested by the PCR assay, the detectable level was 128 pg RNA, a concentration close to one third-stage larva of A.cantonensis, minimum concentration that could be found by naked eyes. The minimum detected total RNA concentration of the third-stage larvae of A.cantonensis was 105 pg by PCR assay.


CONCLUSIONS
A PCR assay has been developed for detecting A.cantonensis larva in Pomacea canaliculata.
Zhang
Yi
Y
National Institute of Parasitic Diseases, Chinese Center for Disease Control and Prevention, WHO Collaborating Center for Malaria, Schistosomiasis and Filariasis, Shanghai 200025, China.
Zhou
Xiao-nong
XN
Liu
He-xiang
HX
Lv
Shan
S
Li
Li-sha
LS
Lin
Jin-xiang
JX
Li
You-song
YS
D017185:Angiostrongylus cantonensis / Q000235:genetics* / Q000254:growth & development; D000818:Animals; D007814:Larva; D018510:RNA, Helminth / Q000235:genetics / Q000302:isolation & purification; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D012908:Snails / Q000469:parasitology*; D017206:Strongylida Infections / Q000175:diagnosis / Q000469:parasitology*
D018510:RNA, Helminth