PCR detection of malaria parasites in desiccated Anopheles mosquitoes is uninhibited by storage time and temperature

Malaria Journal

Research

Joseph Keating¹  Mark A Rider²  Dawn M Wesson²  Kevin A Caillouet³  Brian D Byrd⁴ 

[1] Department of Global Health Systems and Development, Tulane University, 1440 Canal Street, Suite 2200, 70112, New Orleans, LA, USA;Department of Tropical Medicine, Tulane University, 1430 Tulane Avenue, SL-17, 70112, New Orleans, LA, USA;Department of Tropical Medicine, Tulane University, 1430 Tulane Avenue, SL-17, 70112, New Orleans, LA, USA;Department of Biostatistics, Virginia Commonwealth University, 830 East Main Street, 980032, 23298, Richmond, VA, USA;Department of Tropical Medicine, Tulane University, 1430 Tulane Avenue, SL-17, 70112, New Orleans, LA, USA;Environmental Health Sciences Program, Western Carolina University, 156 Joyner Drive, 28723, Cullowhee, NC, USA;

关键词: Plasmodium;    DNA;    Temperature;    Desicant;    Anopheles;    Storage;   

DOI  :  10.1186/1475-2875-11-193

 received in 2012-02-15, accepted in 2012-06-10,  发布年份 2012

来源: Springer

【 摘 要 】

BackgroundReliable methods to preserve mosquito vectors for malaria studies are necessary for detecting Plasmodium parasites. In field settings, however, maintaining a cold chain of storage from the time of collection until laboratory processing, or accessing other reliable means of sample preservation is often logistically impractical or cost prohibitive. As the Plasmodium infection rate of Anopheles mosquitoes is a central component of the entomological inoculation rate and other indicators of transmission intensity, storage conditions that affect pathogen detection may bias malaria surveillance indicators. This study investigated the effect of storage time and temperature on the ability to detect Plasmodium parasites in desiccated Anopheles mosquitoes by real-time polymerase chain reaction (PCR).MethodsLaboratory-infected Anopheles stephensi mosquitoes were chloroform-killed and stored over desiccant for 0, 1, 3, and 6 months while being held at four different temperatures: 28, 37, -20 and -80°C. The detection of Plasmodium DNA was evaluated by real-time PCR amplification of a 111 base pair region of block 4 of the merozoite surface protein.ResultsVarying the storage time and temperature of desiccated mosquitoes did not impact the sensitivity of parasite detection. A two-way factorial analysis of variance suggested that storage time and temperature were not associated with a loss in the ability to detect parasites. Storage of samples at 28°C resulted in a significant increase in the ability to detect parasite DNA, though no other positive associations were observed between the experimental storage treatments and PCR amplification.ConclusionsCold chain maintenance of desiccated mosquito samples is not necessary for real-time PCR detection of parasite DNA. Though field-collected mosquitoes may be subjected to variable conditions prior to molecular processing, the storage of samples over an inexpensive and logistically accessible desiccant will likely ensure accurate assessment of malaria parasite presence without diminishing PCR-detection of parasites in mosquitoes stored for at least six months.

【 授权许可】

CC BY   
© Rider et al.; licensee BioMed Central Ltd. 2012