Abstract
We have developed two reverse transcription polymerase chain reaction (RT-PCR) techniques to detect and quantify the transmission stages (gametocytes) of Plasmodium chabaudi malaria parasites. Both the qualitative and quantitative techniques are based on the amplification of mRNA coding for the P. chabaudi protein Pcs230, which is expressed exclusively in gametocytes. The quantitative RT-PCR (qRT-PCR) technique was developed and validated by examining serial dilutions of known gametocyte densities. The method generated a high correlation between calibration curves of blind samples (R2 = 0.86). The technique was found to be specific, reproducible, and time efficient for quantification of both patent and sub-patent gametocytemia with a sensitivity level 100-1000 times greater than microscopy. The qualitative RT-PCR (RT-PCR) technique was used to monitor the persistence and dynamics of P. chabaudi gametocytes following acute infection. Mice in two independent experiments were sampled for up to 87 days post-infection. RT-PCR showed that gametocytes can persist for up to 8 weeks, post-infection, whereas microscopy could only detect gametocytes up to 6 weeks. Potential applications of the above techniques for studying the ecology, evolution, and epidemiology of malaria transmission are discussed.
| Original language | English |
|---|---|
| Pages (from-to) | 13-20 |
| Number of pages | 8 |
| Journal | Experimental Parasitology |
| Volume | 112 |
| Issue number | 1 |
| DOIs | |
| Publication status | Published - Jan 2006 |
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Keywords
- Chronic infection
- DNA, deoxyribonucleic acid
- dNTPs, deoxyribonucleotide triphosphate mix
- Gametocyte
- Malaria transmission
- pcs230
- Plasmodium chabaudi
- RNA, ribonucleic acid
ASJC Scopus subject areas
- Immunology
- Parasitology
- Infectious Diseases
Cite this
Plasmodium chabaudi : Reverse transcription PCR for the detection and quantification of transmission stage malaria parasites. / Wargo, Andrew R.; Randle, Nadine; Chan, B. H K; Thompson, Joanne; Read, Andrew F.; Babiker, Hamza A.
In: Experimental Parasitology, Vol. 112, No. 1, 01.2006, p. 13-20.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Plasmodium chabaudi
T2 - Reverse transcription PCR for the detection and quantification of transmission stage malaria parasites
AU - Wargo, Andrew R.
AU - Randle, Nadine
AU - Chan, B. H K
AU - Thompson, Joanne
AU - Read, Andrew F.
AU - Babiker, Hamza A.
PY - 2006/1
Y1 - 2006/1
N2 - We have developed two reverse transcription polymerase chain reaction (RT-PCR) techniques to detect and quantify the transmission stages (gametocytes) of Plasmodium chabaudi malaria parasites. Both the qualitative and quantitative techniques are based on the amplification of mRNA coding for the P. chabaudi protein Pcs230, which is expressed exclusively in gametocytes. The quantitative RT-PCR (qRT-PCR) technique was developed and validated by examining serial dilutions of known gametocyte densities. The method generated a high correlation between calibration curves of blind samples (R2 = 0.86). The technique was found to be specific, reproducible, and time efficient for quantification of both patent and sub-patent gametocytemia with a sensitivity level 100-1000 times greater than microscopy. The qualitative RT-PCR (RT-PCR) technique was used to monitor the persistence and dynamics of P. chabaudi gametocytes following acute infection. Mice in two independent experiments were sampled for up to 87 days post-infection. RT-PCR showed that gametocytes can persist for up to 8 weeks, post-infection, whereas microscopy could only detect gametocytes up to 6 weeks. Potential applications of the above techniques for studying the ecology, evolution, and epidemiology of malaria transmission are discussed.
AB - We have developed two reverse transcription polymerase chain reaction (RT-PCR) techniques to detect and quantify the transmission stages (gametocytes) of Plasmodium chabaudi malaria parasites. Both the qualitative and quantitative techniques are based on the amplification of mRNA coding for the P. chabaudi protein Pcs230, which is expressed exclusively in gametocytes. The quantitative RT-PCR (qRT-PCR) technique was developed and validated by examining serial dilutions of known gametocyte densities. The method generated a high correlation between calibration curves of blind samples (R2 = 0.86). The technique was found to be specific, reproducible, and time efficient for quantification of both patent and sub-patent gametocytemia with a sensitivity level 100-1000 times greater than microscopy. The qualitative RT-PCR (RT-PCR) technique was used to monitor the persistence and dynamics of P. chabaudi gametocytes following acute infection. Mice in two independent experiments were sampled for up to 87 days post-infection. RT-PCR showed that gametocytes can persist for up to 8 weeks, post-infection, whereas microscopy could only detect gametocytes up to 6 weeks. Potential applications of the above techniques for studying the ecology, evolution, and epidemiology of malaria transmission are discussed.
KW - Chronic infection
KW - DNA, deoxyribonucleic acid
KW - dNTPs, deoxyribonucleotide triphosphate mix
KW - Gametocyte
KW - Malaria transmission
KW - pcs230
KW - Plasmodium chabaudi
KW - RNA, ribonucleic acid
UR - http://www.scopus.com/inward/record.url?scp=29144442338&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=29144442338&partnerID=8YFLogxK
U2 - 10.1016/j.exppara.2005.08.013
DO - 10.1016/j.exppara.2005.08.013
M3 - Article
VL - 112
SP - 13
EP - 20
JO - Experimental Parasitology
JF - Experimental Parasitology
SN - 0014-4894
IS - 1
ER -