Detection of Schistosoma mansoni in Biomphalaria using nested PCR.
A nested polymerase chain reaction (PCR) protocol was developed for detecting the presence of Schistosoma mansoni sporocysts in intermediate host snails of the genus Biomphalaria. To accomplish this, rDNA genes encoding the 18S rRNA of S. mansoni and Biomphalaria alexandrina from Egypt were sequenced, as were 18S-encoding genes of the 13-16-R1 and Salvador strains of Biomphalaria glabrata. Based on a comparison of host and parasite sequences, a nested set of PCR primers was designed to allow specific amplification of portions of S. mansoni 18S rDNA. These primers allowed detection of as little as 10 fg of S. mansoni DNA diluted in 100 ng of snail DNA and did not allow amplification of snail 18S sequences. Using nested PCR, the presence of a single S. mansoni sporocyst within an adult snail could be detected at 1 day postexposure. In DNA samples extracted from each of 74 snails of the M-line strain of B. glabrata exposed to from 1 to 10 S. mansoni miracidia for intervals ranging from 1 to 44 days, use of the outside primer pair alone detected the parasite's presence in 51% of the snails, whereas the sequential use of outside and nested primer pairs detected parasites in 92% of the snails. This approach has utility in determining if snails in endemic areas bear prepatent or inactive infections and in assessing the degree of compatibility between local snail and schistosome populations. It will also facilitate studies of resistance of snails to infection.
Hanelt
B
B
Department of Biology, University of New Mexico, Albuquerque 87131, USA.
Adema
C M
CM
Mansour
M H
MH
Loker
E S
ES
D000818:Animals; D001483:Base Sequence; D001700:Biomphalaria / Q000469:parasitology*; D003001:Cloning, Molecular; D017931:DNA Primers / Q000737:chemistry / Q000235:genetics; D018509:DNA, Helminth / Q000032:analysis / Q000737:chemistry; D004275:DNA, Ribosomal / Q000032:analysis* / Q000737:chemistry; D005260:Female; D008297:Male; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D018510:RNA, Helminth / Q000235:genetics; D012337:RNA, Ribosomal, 18S / Q000235:genetics*; D012550:Schistosoma mansoni / Q000235:genetics / Q000302:isolation & purification*; D012680:Sensitivity and Specificity; D016415:Sequence Alignment
D017931:DNA Primers; D018509:DNA, Helminth; D004275:DNA, Ribosomal; D018510:RNA, Helminth; D012337:RNA, Ribosomal, 18S