An Optimized Real-Time qPCR Method for the Effective Detection of Human Malaria Infections
Polymerase chain reaction, although an expensive method for the detection of human <i>Plasmodium</i> spp., is still considered the finest for the diagnosis of malaria. The conventional diagnostic PCR is an inexpensive process but consumes a lot of time, reagents and lacks sensitivity. On...
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| Format: | Article |
| Language: | English |
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MDPI AG
2021-04-01
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| Series: | Diagnostics |
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| Online Access: | https://www.mdpi.com/2075-4418/11/5/736 |
| _version_ | 1797536775250378752 |
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| author | Saiful Arefeen Sazed Mohammad Golam Kibria Mohammad Shafiul Alam |
| author_facet | Saiful Arefeen Sazed Mohammad Golam Kibria Mohammad Shafiul Alam |
| author_sort | Saiful Arefeen Sazed |
| collection | DOAJ |
| description | Polymerase chain reaction, although an expensive method for the detection of human <i>Plasmodium</i> spp., is still considered the finest for the diagnosis of malaria. The conventional diagnostic PCR is an inexpensive process but consumes a lot of time, reagents and lacks sensitivity. On the other hand, real-time PCR assays currently being used are mostly probe-based expensive methods and sometimes not feasible to detect all the species in a single amplification reaction condition. Here we have established a real-time PCR method that is time and cost effective with a single protocol to detect and distinguish five human <i>Plasmodium</i> species using the existing primers efficiently. The primers used here are being used in the conventional method and the sensitivity as well as specificity of this method has also been immensely improved (100%). The lower limit of detection for <i>Plasmodium falciparum</i>, <i>Plasmodium vivax</i> and <i>Plasmodium malariae</i> are 0.064 parasites/µL, 1.6 parasites/µL, and 0.32 parasites/µL respectively and no cross reactivity was observed. Besides, we have analyzed melt curves that can be used for further species confirmation and validation purposes using multiplex systems. This method, therefore, can be considered as an alternative to the existing lineup for molecular diagnosis of malaria in endemic countries. |
| first_indexed | 2024-03-10T12:05:35Z |
| format | Article |
| id | doaj.art-45b95f37eb0441ecaab930c68941af83 |
| institution | Directory Open Access Journal |
| issn | 2075-4418 |
| language | English |
| last_indexed | 2024-03-10T12:05:35Z |
| publishDate | 2021-04-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Diagnostics |
| spelling | doaj.art-45b95f37eb0441ecaab930c68941af832023-11-21T16:34:57ZengMDPI AGDiagnostics2075-44182021-04-0111573610.3390/diagnostics11050736An Optimized Real-Time qPCR Method for the Effective Detection of Human Malaria InfectionsSaiful Arefeen Sazed0Mohammad Golam Kibria1Mohammad Shafiul Alam2Infectious Diseases Division, International Centre for Diarrheal Diseases Research, Bangladesh Mohakhali, Dhaka 1212, BangladeshInfectious Diseases Division, International Centre for Diarrheal Diseases Research, Bangladesh Mohakhali, Dhaka 1212, BangladeshInfectious Diseases Division, International Centre for Diarrheal Diseases Research, Bangladesh Mohakhali, Dhaka 1212, BangladeshPolymerase chain reaction, although an expensive method for the detection of human <i>Plasmodium</i> spp., is still considered the finest for the diagnosis of malaria. The conventional diagnostic PCR is an inexpensive process but consumes a lot of time, reagents and lacks sensitivity. On the other hand, real-time PCR assays currently being used are mostly probe-based expensive methods and sometimes not feasible to detect all the species in a single amplification reaction condition. Here we have established a real-time PCR method that is time and cost effective with a single protocol to detect and distinguish five human <i>Plasmodium</i> species using the existing primers efficiently. The primers used here are being used in the conventional method and the sensitivity as well as specificity of this method has also been immensely improved (100%). The lower limit of detection for <i>Plasmodium falciparum</i>, <i>Plasmodium vivax</i> and <i>Plasmodium malariae</i> are 0.064 parasites/µL, 1.6 parasites/µL, and 0.32 parasites/µL respectively and no cross reactivity was observed. Besides, we have analyzed melt curves that can be used for further species confirmation and validation purposes using multiplex systems. This method, therefore, can be considered as an alternative to the existing lineup for molecular diagnosis of malaria in endemic countries.https://www.mdpi.com/2075-4418/11/5/736<i>Plasmodium</i>malariareal time PCRasymptomaticdiagnosisnested PCR |
| spellingShingle | Saiful Arefeen Sazed Mohammad Golam Kibria Mohammad Shafiul Alam An Optimized Real-Time qPCR Method for the Effective Detection of Human Malaria Infections Diagnostics <i>Plasmodium</i> malaria real time PCR asymptomatic diagnosis nested PCR |
| title | An Optimized Real-Time qPCR Method for the Effective Detection of Human Malaria Infections |
| title_full | An Optimized Real-Time qPCR Method for the Effective Detection of Human Malaria Infections |
| title_fullStr | An Optimized Real-Time qPCR Method for the Effective Detection of Human Malaria Infections |
| title_full_unstemmed | An Optimized Real-Time qPCR Method for the Effective Detection of Human Malaria Infections |
| title_short | An Optimized Real-Time qPCR Method for the Effective Detection of Human Malaria Infections |
| title_sort | optimized real time qpcr method for the effective detection of human malaria infections |
| topic | <i>Plasmodium</i> malaria real time PCR asymptomatic diagnosis nested PCR |
| url | https://www.mdpi.com/2075-4418/11/5/736 |
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