Volume 78, Issue 3
  • ISSN: 0002-9637
  • E-ISSN: 1476-1645


We developed real-time fluorescence resonance energy transfer (FRET) polymerase chain reaction (PCR) combined with melting curve analysis for detection of DNA in blood-fed mosquitoes. Real-time FRET PCR is based on a fluorescence melting curve analysis of hybrid formed between amplicons generated from a family of repeated DNA element, 153-bp I repeated sequence, specific to genus and specific fluorophore-labeled probes. The –infected mosquitoes were differentiated from –infected and uninfected mosquitoes and from genomic DNA of – and –infected human red blood cells and human leukocytes by their melting temperature. Sensitivity and specificity were both 100%. Melting curve analysis produces a rapid, accurate, and sensitive alternative for specific detection of in mosquitoes, allows high throughput, and can be performed on small samples. This method has the potential for endemic area mapping or monitoring effect of brugian filariasis mass treatment programs.


Article metrics loading...

Loading full text...

Full text loading...



  1. World Health Organization, 1992. Lymphatic filariasis: the disease and its control. Fifth report of the WHO Expert Committee on Filariasis. World Health Organ Tech Rep Ser 821 : 1–71.
  2. Weil GJ, Ramzy RM, 2007. Diagnostic tools for filariasis elimination programs. Trends Parasitol 23 : 78–82.
  3. Rao RU, Weil GJ, Fischer K, Supali T, Fischer P, 2006. Detection of Brugia parasite DNA in human blood by real-time PCR. J Clin Microbiol 44 : 3887–3893.
  4. Williams SA, Laney SJ, Bierwert LA, Saunders LJ, Boakye DA, Fischer P, Goodman D, Helmy H, Hoti SL, Vasuki V, Lammie PJ, Plichart C, Ramzy RM, Ottesen EA, 2002. Development and standardization of a rapid, PCR-based method for the detection of Wuchereria bancrofti in mosquitoes, for xenomonitoring the human prevalence of bancroftian filariasis. Ann Trop Med Parasitol 96 (Suppl): S41–S46.
  5. McCarthy JS, Zhong M, Gopinath R, Ottesen EA, Williams SA, Nutman TB, 1996. Evaluation of a polymerase chain reaction-based assay for diagnosis of Wuchereria bancrofti infection. J Infect Dis 173 : 1510–1514.
  6. Ramzy RM, Farid HA, Kamal IH, Ibrahim GH, Morsy ZS, Faris R, Weil GJ, Williams SA, Gad AM, 1997. A polymerase chain reaction-based assay for detection of Wuchereria bancrofti in human blood and Culex pipiens. Trans R Soc Trop Med Hyg 91 : 156–160.
  7. Zarlenga DS, Higgins J, 2001. PCR as a diagnostic and quantitative technique in veterinary parasitology. Vet Parasitol 101 : 215–230.
  8. Menard A, Dachet F, Prouzet-Mauleon V, Oleastro M, Megraud F, 2005. Development of a real-time fluorescence resonance energy transfer PCR to identify the main pathogenic Campylobacter spp. Clin Microbiol Infect 11 : 281–287.
  9. Hakhverdyan M, Rasmussen TB, Thoren P, Uttenthal A, Belak S, 2006. Development of a real-time PCR assay based on primer-probe energy transfer for the detection of swine vesicular disease virus. Arch Virol 151 : 2365–2376.
  10. Abdelbaqi K, Buissonniere A, Prouzet-Mauleon V, Gresser J, Wesley I, Megraud F, Menard A, 2007. Development of a real-time fluorescence resonance energy transfer PCR to detect arcobacter species. J Clin Microbiol 45 : 3015–3021.
  11. Walker NJ, 2001. Real-time and quantitative PCR: applications to mechanism-based toxicology. J Biochem Mol Toxicol 15 : 121–127.
  12. Rao RU, Atkinson LJ, Ramzy RM, Helmy H, Farid HA, Bockarie MJ, Susapu M, Laney SJ, Williams SA, Weil GJ, 2006. A real-time PCR-based assay for detection of Wuchereria bancrofti DNA in blood and mosquitoes. Am J Trop Med Hyg 74 : 826–832.
  13. Fischer P, Erickson SM, Fischer K, Fuchs JF, Rao RU, Christensen BM, Weil GJ, 2007. Persistence of Brugia malayi DNA in vector and non-vector mosquitoes: implications for xenomonitoring and transmission monitoring of lymphatic filariasis. Am J Trop Med Hyg 76 : 502–507.
  14. Lulitanond V, Intapan PM, Pipitgool V, Choochote W, Maleewong W, 2004. Rapid detection of Wuchereria bancrofti in mosquitoes by LightCycler polymerase chain reaction and melting curve analysis. Parasitol Res 94 : 337–341.
  15. McReynolds LA, DeSimone SM, Williams SA, 1986. Cloning and comparison of repeated DNA sequences from the human filarial parasite Brugia malayi and the animal parasite Brugia pahangi. Proc Natl Acad Sci U S A 83 : 797–801.
  16. Choochote W, Keha P, Sukhavat K, Khamboonruang C, Sukontason K, 1987. Aedes (Finlaya) togoi Theobald 1907, Chanthaburi strain, a laboratory vector in studies of filariasis in Thailand. Southeast Asian J Trop Med Public Health 18 : 259–260.
  17. Maleewong W, Choochote W, Sukhavat K, Khamboonruang C, Arunyanart C, 1987. Scanning electron microscopic study of third-stage larva of Wuchereria bancrofti and Brugia malayi in Thailand. Southeast Asian J Trop Med Public Health 18 : 261–264.
  18. Chomcharn Y, Surathin K, Bunnag D, Sucharit S, Harinasuta T, 1980. Effect of a single dose of primaquine on a Thai strain of Plasmodium falciparum. Southeast Asian J Trop Med Public Health 18 : 408–412.
  19. Ghedin E, Wang S, Foster JM, Slatko BE, 2004. First sequenced genome of a parasitic nematode. Trends Parasitol 20 : 151–153.
  20. Fischer P, Wibowo H, Pischke S, Ruckert P, Liebau E, Ismid IS, Supali T, 2002. PCR-based detection and identification of the filarial parasite Brugia timori from Alor Island, Indonesia. Ann Trop Med Parasitol 96 : 809–821.
  21. Fischer P, Boakye D, Hamburger J, 2003. Polymerase chain reaction-based detection of lymphatic filariasis. Med Microbiol Immunol (Berl) 192 : 3–7.

Data & Media loading...

  • Received : 10 Oct 2007
  • Accepted : 08 Dec 2007

Most Cited This Month

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error