Nonradioactive heteroduplex tracking assay for the detection of minority-variant chloroquine-resistant <it>Plasmodium falciparum </it>in Madagascar
<p>Abstract</p> <p>Background</p> <p>Strains of <it>Plasmodium falciparum </it>genetically resistant to chloroquine (CQ) due to the presence of <it>pfcrt </it>76T appear to have been recently introduced to the island of Madagascar. The prevalence...
| Main Authors: | , , , , , |
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| Format: | Article |
| Language: | English |
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BMC
2009-03-01
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| Series: | Malaria Journal |
| Online Access: | http://www.malariajournal.com/content/8/1/47 |
| _version_ | 1818146740654571520 |
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| author | Mwapasa Victor Ariey Frédéric Ramarosandratana Benjamin Randrianarivelojosia Milijaona Juliano Jonathan J Meshnick Steven R |
| author_facet | Mwapasa Victor Ariey Frédéric Ramarosandratana Benjamin Randrianarivelojosia Milijaona Juliano Jonathan J Meshnick Steven R |
| author_sort | Mwapasa Victor |
| collection | DOAJ |
| description | <p>Abstract</p> <p>Background</p> <p>Strains of <it>Plasmodium falciparum </it>genetically resistant to chloroquine (CQ) due to the presence of <it>pfcrt </it>76T appear to have been recently introduced to the island of Madagascar. The prevalence of such resistant genotypes is reported to be low (< 3%) when evaluated by conventional PCR. However, these methods are insensitive to low levels of mutant parasites present in patients with polyclonal infections. Thus, the current estimates may be an under representation of the prevalence of the CQ-resistant <it>P. falciparum </it>isolates on the island. Previously, minority variant chloroquine resistant parasites were described in Malawian patients using an isotopic heteroduplex tracking assay (HTA), which can detect <it>pfcrt </it>76T-bearing <it>P. falciparum </it>minority variants in individual patients that were undetectable by conventional PCR. However, as this assay required a radiolabeled probe, it could not be used in many resource-limited settings.</p> <p>Methods</p> <p>This study describes a digoxigenin (DIG)-labeled chemiluminescent heteroduplex tracking assay (DIG-HTA) to detect <it>pfcrt </it>76T-bearing minority variant <it>P. falciparum</it>. This assay was compared to restriction fragment length polymorphism (RFLP) analysis and to the isotopic HTA for detection of genetically CQ-resistant parasites in clinical samples.</p> <p>Results</p> <p>Thirty one clinical <it>P. falciparum </it>isolates (15 primary isolates and 16 recurrent isolates) from 17 Malagasy children treated with CQ for uncomplicated malaria were genotyped for the <it>pfcrt </it>K76T mutation. Two (11.7%) of 17 patients harboured genetically CQ-resistant <it>P. falciparum </it>strains after therapy as detected by HTA. RFLP analysis failed to detect any <it>pfcrt </it>K76T-bearing isolates.</p> <p>Conclusion</p> <p>These findings indicate that genetically CQ-resistant <it>P. falciparum </it>are more common than previously thought in Madagascar even though the fitness of the minority variant <it>pfcrt </it>76T parasites remains unclear. In addition, HTAs for malaria drug resistance alleles are promising tools for the surveillance of anti-malarial resistance. The use of a non-radioactive label allows for the use of HTAs in malaria endemic countries.</p> |
| first_indexed | 2024-12-11T12:24:10Z |
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| institution | Directory Open Access Journal |
| issn | 1475-2875 |
| language | English |
| last_indexed | 2024-12-11T12:24:10Z |
| publishDate | 2009-03-01 |
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| spelling | doaj.art-33bc0f27b93c4b73adaca334c30224eb2022-12-22T01:07:26ZengBMCMalaria Journal1475-28752009-03-01814710.1186/1475-2875-8-47Nonradioactive heteroduplex tracking assay for the detection of minority-variant chloroquine-resistant <it>Plasmodium falciparum </it>in MadagascarMwapasa VictorAriey FrédéricRamarosandratana BenjaminRandrianarivelojosia MilijaonaJuliano Jonathan JMeshnick Steven R<p>Abstract</p> <p>Background</p> <p>Strains of <it>Plasmodium falciparum </it>genetically resistant to chloroquine (CQ) due to the presence of <it>pfcrt </it>76T appear to have been recently introduced to the island of Madagascar. The prevalence of such resistant genotypes is reported to be low (< 3%) when evaluated by conventional PCR. However, these methods are insensitive to low levels of mutant parasites present in patients with polyclonal infections. Thus, the current estimates may be an under representation of the prevalence of the CQ-resistant <it>P. falciparum </it>isolates on the island. Previously, minority variant chloroquine resistant parasites were described in Malawian patients using an isotopic heteroduplex tracking assay (HTA), which can detect <it>pfcrt </it>76T-bearing <it>P. falciparum </it>minority variants in individual patients that were undetectable by conventional PCR. However, as this assay required a radiolabeled probe, it could not be used in many resource-limited settings.</p> <p>Methods</p> <p>This study describes a digoxigenin (DIG)-labeled chemiluminescent heteroduplex tracking assay (DIG-HTA) to detect <it>pfcrt </it>76T-bearing minority variant <it>P. falciparum</it>. This assay was compared to restriction fragment length polymorphism (RFLP) analysis and to the isotopic HTA for detection of genetically CQ-resistant parasites in clinical samples.</p> <p>Results</p> <p>Thirty one clinical <it>P. falciparum </it>isolates (15 primary isolates and 16 recurrent isolates) from 17 Malagasy children treated with CQ for uncomplicated malaria were genotyped for the <it>pfcrt </it>K76T mutation. Two (11.7%) of 17 patients harboured genetically CQ-resistant <it>P. falciparum </it>strains after therapy as detected by HTA. RFLP analysis failed to detect any <it>pfcrt </it>K76T-bearing isolates.</p> <p>Conclusion</p> <p>These findings indicate that genetically CQ-resistant <it>P. falciparum </it>are more common than previously thought in Madagascar even though the fitness of the minority variant <it>pfcrt </it>76T parasites remains unclear. In addition, HTAs for malaria drug resistance alleles are promising tools for the surveillance of anti-malarial resistance. The use of a non-radioactive label allows for the use of HTAs in malaria endemic countries.</p>http://www.malariajournal.com/content/8/1/47 |
| spellingShingle | Mwapasa Victor Ariey Frédéric Ramarosandratana Benjamin Randrianarivelojosia Milijaona Juliano Jonathan J Meshnick Steven R Nonradioactive heteroduplex tracking assay for the detection of minority-variant chloroquine-resistant <it>Plasmodium falciparum </it>in Madagascar Malaria Journal |
| title | Nonradioactive heteroduplex tracking assay for the detection of minority-variant chloroquine-resistant <it>Plasmodium falciparum </it>in Madagascar |
| title_full | Nonradioactive heteroduplex tracking assay for the detection of minority-variant chloroquine-resistant <it>Plasmodium falciparum </it>in Madagascar |
| title_fullStr | Nonradioactive heteroduplex tracking assay for the detection of minority-variant chloroquine-resistant <it>Plasmodium falciparum </it>in Madagascar |
| title_full_unstemmed | Nonradioactive heteroduplex tracking assay for the detection of minority-variant chloroquine-resistant <it>Plasmodium falciparum </it>in Madagascar |
| title_short | Nonradioactive heteroduplex tracking assay for the detection of minority-variant chloroquine-resistant <it>Plasmodium falciparum </it>in Madagascar |
| title_sort | nonradioactive heteroduplex tracking assay for the detection of minority variant chloroquine resistant it plasmodium falciparum it in madagascar |
| url | http://www.malariajournal.com/content/8/1/47 |
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