A simple and rapid DNA extraction method for the detection of Wuchereria bancrofti infection in the vector mosquito, Culex quinquefasciatus by Ssp I PCR assay.
A simple, rapid and inexpensive method for the extraction of DNA from filarial vector, Culex quinquefasciatus, useful in Ssp I PCR assay for xenomonitoring of infection with Wuchereria bancrofti is presented. The DNA extracted by this method was found suitable for PCR detection of W. bancrofti infection in pools of 10-30 mosquitoes. The PCR assay employing the simplified DNA extraction method was evaluated for its sensitivity on field caught Cx. quinquefasciatus, in comparison with the conventional dissection and microscopy technique. When assayed on dissection washings of vector mosquitoes the PCR assay detected 45 pools out of 49 dissection positive pools as positive for infection and hence found to be less sensitive than the conventional technique. The reason for detecting four dissection positive pools as negatives by the PCR assay may be due to the loss of a few numbers of parasites (1-3) present in these pools during the transfer of washings of dissected mosquitoes. The PCR assay detected ten out of 72 dissection negative pools as positives, while it did not detect any of the 62 known negative (laboratory reared, uninfected) mosquito pools as positives. When 38 pools (10 mosquitoes/pool) of intact mosquitoes were assessed for infection by each method, the infection rates obtained by the two methods were almost similar (3.35 and 3.01%, respectively, for conventional method and PCR assay). The results thus show that the DNA extraction method, which is simple, rapid, safe and inexpensive, is efficient to generate DNA from vector mosquitoes useful in PCR assay and hence has potential application in xenomonitoring.
Vasuki
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Vector Control Research Centre (ICMR), Medical Complex, Indira Nagar, 600 006 Pondicherry, India. vvasuki@yahoo.com
Hoti
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Sadanandane
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Jambulingam
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D000818:Animals; D003465:Culex / Q000235:genetics / Q000469:parasitology*; D018509:DNA, Helminth / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D005260:Female; D005368:Filariasis / Q000469:parasitology*; D006801:Humans; D007303:Insect Vectors / Q000469:parasitology*; D018895:Microsatellite Repeats / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D014958:Wuchereria bancrofti / Q000235:genetics*
D018509:DNA, Helminth