Genetic Polymorphisms Influence
            <i>Plasmodium ovale</i>
            PCR Detection Accuracy

Calderaro, Adriana; Piccolo, Giovanna; Perandin, Francesca; Gorrini, Chiara; Peruzzi, Simona; Zuelli, C.; Ricci, Lucila Costallat; Manca, N.; Dettori, G.; Chezzi, Carlo; Snounou, Georges

doi:10.1128/jcm.02316-06

Cited by 99 publications

(127 citation statements)

References 30 publications

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“…One of the major limitations of the Rougemont method is the ability to detect mixed infections (3,4,19). This is likely due to competition of the conserved primers for the different templates and favors amplification of the species with the highest level of infection.…”

Section: Resultsmentioning

confidence: 99%

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Multiplexed Real-Time PCR Assay for Discrimination of Plasmodium Species with Improved Sensitivity for Mixed Infections

Shokoples

Ndao

Kowalewska-Grochowska

et al. 2009

J Clin Microbiol

228

260

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The implementation of real-time PCR for the diagnosis of malaria has been hampered by poor sensitivity for the detection of mixed infections. We have optimized a method that enhances the sensitivity of detection of minor species in mixed infections within a single multiplex reaction. Our assay uses species-specific forward primers in combination with a conserved reverse primer and largely overcomes primer competition for the minor species DNA. With a blind panel of clinical samples, we successfully identified the species in 13/16 mixed infections. This assay was further validated with 91 blood samples and demonstrated a specificity and sensitivity for single infections of 100% compared with nested PCR as the "gold standard." This test has been implemented for routine confirmation of malaria species in Alberta, Canada. In comparison with species identification by microscopy, the real-time PCR test demonstrated greater sensitivity for the identification of species causing low-level and mixed infections and for the discrimination of Plasmodium species other than Plasmodium falciparum. Our experience supports a role for real-time PCR in the identification of malarial species in conjunction with microscopy.

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Section: Resultsmentioning

confidence: 99%

“…One method uses primer sets specific to P. falciparum, P. vivax, and P. ovale but does not include primers to detect P. malariae (18). This method also fails to capture DNA sequence variations in circulating strains of P. ovale (3,4).…”

mentioning

confidence: 99%

Multiplexed Real-Time PCR Assay for Discrimination of Plasmodium Species with Improved Sensitivity for Mixed Infections

Shokoples

Ndao

Kowalewska-Grochowska

et al. 2009

J Clin Microbiol

228

260

View full text Add to dashboard Cite

show abstract

“…This procedure is cheap and simple, but is a labor-intensive procedure, which requires well-trained personnel (17). When parasitemia is very low, the data of microscopy diagnosis are limited, and in some cases biased by the inability to devote the necessary amount of time to the examination of blood smears (17).…”

Section: Discussionmentioning

confidence: 99%

The Incidence of Current Infection with Different Human Malaria Species by Polymerase Chain Reaction for Diagnosis of Suspicious Malaria Patients on Elimination Region Sistan and Baluchistan Province, Southeast of Iran

Ebrahimzadeh

Dalir

Mehravaran

et al. 2017

Jundishapur J Microbiol

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Background: Malaria, as a parasitic disease, is one of the most important public health problems in Iran. Malaria is mainly diagnosed by peripheral blood smear, stained by Giemsa; in Iran it is also diagnosed by blood smear that is highly depends on technician's skills and laboratory properties. Objectives: Correct diagnosis of malaria and identification of human malaria species in spite of the measures taken to eliminate the disease in Iran, have made the complete understanding of malaria epidemiology critical. Therefore, the current study aimed at investigating the epidemiology of 2 species of human Plasmodium in Sistan and Baluchistan province using polymerase chain reaction (PCR) method. Methods: The present descriptive study was conducted on 100 patients suspected to malaria infection who referred to health centers of Chabahar, Iranshahr, Nikshahr, and Sarbaz districts. DNAs were extracted from blood samples using the specific kit, and nested-PCR reaction was performed to identify the Plasmodium species according to NP-2013 protocol. Results: Molecular analysis was performed on 100 samples suspected of malaria; 84 negative and 16 positive samples were detected including 8 Plasmodium vivax, 2 P. falciparum, and 6 mixed infections (P. vivax and P. falciparum). No P. ovale or P. malariae was observed. Conclusions:The results showed that malaria had a decreasing trend in Sistan and Baluchistan province. Therefore, the malaria elimination program is applicable and attainable in this region as a goal.

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“…For example, several assays only distinguish P. falciparum from other Plasmodium species (12,13). One assay cannot detect P. malariae (14) and additionally could miss variants of P. ovale (15,16). Clinical specificity is not ensured, since blood samples from patients with P. ovale and P. malariae are difficult to obtain and infrequently tested.…”

Section: Figmentioning

confidence: 99%

Multiplex 5′ Nuclease Quantitative Real-Time PCR for Clinical Diagnosis of Malaria and Species-Level Identification and Epidemiologic Evaluation of Malaria-Causing Parasites, Including Plasmodium knowlesi

et al. 2013

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Molecular diagnosis of malaria offers many potential advantages over microscopy, including identification of malaria to the species level in an era with few experienced microscopists. We developed high-throughput multiplex 5= nuclease quantitative PCR (qPCR) assays, with the potential to support large studies, to specifically identify Plasmodium falciparum, P. vivax, P. ovale, P. malariae, and P. knowlesi. We compared qPCR to microscopy and confirmed discordant results with an alternative target PCR assay. The assays specifically detected 1 to 6 parasites/l of blood. The clinical sensitivities (95% confidence intervals [CIs]) of the 4-plex assay to detect microscopically confirmed malaria were 95.8% (88.3 to 99.1%) for P. falciparum, 89.5% (75.2 to 97.1%) for P. vivax, 94.1% (71.3 to 99.9%) for P. ovale, and 100% (66.4 to 100%) for P. malariae. The specificities (95% CIs) were 98.6% (92.4 to 100%) for P. falciparum, 99% (84.8 to 100%) for P. vivax, 98.4% (94.4 to 99.8%) for P. ovale, and 99.3% (95.9 to 100%) for P. malariae. The clinical specificity for samples without malaria was 100%. The clinical sensitivity of the 5-plex assay for confirmed P. knowlesi malaria was 100% (95% CI, 69.2 to 100%), and the clinical specificity was 100% (95% CI, 87.2 to 100%). Coded retesting and testing with an alternative target PCR assay showed improved sensitivity and specificity of multiplex qPCR versus microscopy. Additionally, 91.7% (11/12) of the samples with uncertain species by microscopy were identified to the species level identically by both our multiplex qPCR assay and the alternative target PCR assay, including 9 P. falciparum infections. Multiplex qPCR can rapidly and simultaneously identify all 5 Plasmodium species known to cause malaria in humans, and it offers an alternative or adjunct to microscopy for clinical diagnosis as well as a needed high-throughput tool for research.

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Genetic Polymorphisms Influence Plasmodium ovale PCR Detection Accuracy

Cited by 99 publications

References 30 publications

Multiplexed Real-Time PCR Assay for Discrimination of Plasmodium Species with Improved Sensitivity for Mixed Infections

Multiplexed Real-Time PCR Assay for Discrimination of Plasmodium Species with Improved Sensitivity for Mixed Infections

The Incidence of Current Infection with Different Human Malaria Species by Polymerase Chain Reaction for Diagnosis of Suspicious Malaria Patients on Elimination Region Sistan and Baluchistan Province, Southeast of Iran

Multiplex 5′ Nuclease Quantitative Real-Time PCR for Clinical Diagnosis of Malaria and Species-Level Identification and Epidemiologic Evaluation of Malaria-Causing Parasites, Including Plasmodium knowlesi

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