Real-time PCR assay and rapid diagnostic tests for the diagnosis of clinically suspected malaria patients in Bangladesh

Alam, Mohammad Shafiul; Mohon, Abu Naser; Mustafa, Shariar; Khan, Wasif Ali; Islam, Nazrul; Karim, Mohammad Jahirul; Khanum, Hamida; Sullivan, David J.; Haque, Rashidul

doi:10.1186/1475-2875-10-175

Cited by 59 publications

(71 citation statements)

References 28 publications

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“…Microscopy results were confirmed by both real time PCR and nested PCR [18, 19]. If any discrepancies were found by different diagnostic methods, those samples were excluded from the study.…”

Section: Methodsmentioning

confidence: 99%

Mutations in Plasmodium falciparum K13 propeller gene from Bangladesh (2009–2013)

Mohon

Alam

Bayih

et al. 2014

Malar J

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BackgroundBangladesh is a malaria hypo-endemic country sharing borders with India and Myanmar. Artemisinin combination therapy (ACT) remains successful in Bangladesh. An increase of artemisinin-resistant malaria parasites on the Thai-Cambodia and Thai-Myanmar borders is worrisome. K13 propeller gene (PF3D7_1343700 or PF13_0238) mutations have been linked to both in vitro artemisinin resistance and in vivo slow parasite clearance rates. This group undertook to evaluate if mutations seen in Cambodia have emerged in Bangladesh where ACT use is now standard for a decade.MethodsSamples were obtained from Plasmodium falciparum-infected malaria patients from Upazila health complexes (UHC) between 2009 and 2013 in seven endemic districts of Bangladesh. These districts included Khagrachari (Matiranga UHC), Rangamati (Rajasthali UHC), Cox’s Bazar (Ramu and Ukhia UHC), Bandarban (Lama UHC), Mymensingh (Haluaghat UHC), Netrokona (Durgapur and Kalmakanda UHC), and Moulvibazar (Sreemangal and Kamalganj UHC).ResultsOut of 296 microscopically positive P. falciparum samples, 271 (91.6%) were confirmed as mono-infections by both real-time PCR and nested PCR. The K13 propeller gene from 253 (93.4%) samples was sequenced bi-directionally. One non-synonymous mutation (A578S) was found in Bangladeshi clinical isolates. The A578S mutation was confirmed and lies adjacent to the C580Y mutation, the major mutation causing delayed parasite clearance in Cambodia. Based on computational modeling A578S should have a significant effect on tertiary structure of the protein.ConclusionThe data suggest that P. falciparum in Bangladesh remains free of the C580Y mutation linked to delayed parasite clearance. However, the mutation A578S is present and based on structural analysis could affect K13 gene function. Further in vivo clinical studies are required to validate the effect of this mutation.

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Section: Methodsmentioning

confidence: 99%

Mutations in Plasmodium falciparum K13 propeller gene from Bangladesh (2009–2013)

Mohon

Alam

Bayih

et al. 2014

Malar J

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“…The sensitivity and specificity of qPCR for P. falciparum was 97.1 and 97.6%, respectively, while for P. vivax 95.2 and 98.1% [6]. Any mixed ( P. falciparum and P. vivax ) infection diagnosed by qPCR was not considered in this study.…”

Section: Methodsmentioning

confidence: 99%

Performance of a HRP-2/pLDH based rapid diagnostic test at the Bangladesh-India-Myanmar border areas for diagnosis of clinical malaria

Elahi

Mohon

Khan

et al. 2013

Malar J

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BackgroundThe rapid diagnostic test (RDT) has been adopted in contemporary malaria control and management programmes around the world as it represents a fast and apt alternative for malaria diagnosis in a resource-limited setting. This study assessed the performance of a HRP-2/pLDH based RDT (Parascreen® Pan/Pf) in a laboratory setting utilizing clinical samples obtained from the field.MethodsWhole blood samples were obtained from febrile patients referred for malaria diagnosis by clinicians from two different Upazila Health Complexes (UHCs) located near the Bangladesh-India and Bangladesh-Myanmar border where malaria is endemic. RDT was performed on archived samples and sensitivity and specificity evaluated with expert microscopy (EM) and quantitative PCR (qPCR).ResultsA total of 327 clinical samples were made available for the study, of which 153 were Plasmodium falciparum-positive and 54 were Plasmodium vivax-positive. In comparison with EM, for P. falciparum malaria, the RDT had sensitivity: 96.0% (95% CI, 91.2-98.3) and specificity: 98.2% (95% CI, 94.6-99.5) and for P. vivax, sensitivity: 90.7% (95% CI, 78.9-96.5) and specificity: 98.9% (95% CI, 96.5-99.7). Comparison with qPCR showed, for P. falciparum malaria, sensitivity: 95.4% (95% CI, 90.5-98.0) and specificity: 98.8% (95% CI, 95.4-99.7) and for P. vivax malaria, sensitivity: 89.0% (95% CI,77.0-95.4) and specificity: 98.8% (95% CI, 96.5-99.7). Sensitivity varied according to different parasitaemia for falciparum and vivax malaria diagnosis.ConclusionParascreen® Pan/Pf Rapid test for malaria showed acceptable sensitivity and specificity in border belt endemic areas of Bangladesh when compared with EM and qPCR.

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“…For areas where there are high rates of transmission of P. vivax or other Plasmodium species, additional species-specific detection lines should be added to the db-PCR-NALFIA detection device. Since currently available RDTs have a low sensitivity for the detection of P. vivax (1,10), in remote field settings microscopy is still the only relatively easy, although very time-consuming, field-deployable method for the detection of P. vivax (10). The current db-PCR-NALFIA consists of a combination of components from two commercially available kits, the buffer component of the Phusion direct blood PCR kit and the separately available Phire Hotstart II DNA polymerase.…”

Section: Discussionmentioning

confidence: 99%

Direct Blood PCR in Combination with Nucleic Acid Lateral Flow Immunoassay for Detection of Plasmodium Species in Settings Where Malaria Is Endemic

et al. 2012

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fDeclining malaria transmission and known difficulties with current diagnostic tools for malaria, such as microscopy and rapid diagnostic tests (RDTs) in particular at low parasite densities, still warrant the search for sensitive diagnostic tests. Molecular tests need substantial simplification before implementation in clinical settings in countries where malaria is endemic. Direct blood PCR (db-PCR), circumventing DNA extraction, to detect Plasmodium was developed and adapted to be visualized by nucleic acid lateral flow immunoassay (NALFIA). The assay was evaluated in the laboratory against samples from confirmed Sudanese patients (n ‫؍‬ 51), returning travelers (n ‫؍‬ 214), samples from the Dutch Blood Bank (n ‫؍‬ 100), and in the field in Burkina Faso (n ‫؍‬ 283) and Thailand (n ‫؍‬ 381) on suspected malaria cases and compared to RDT and microscopy. The sensitivity and specificity of db-PCR-NALFIA compared to the initial diagnosis in the laboratory were 94.4% (95% confidence interval [CI] ‫؍‬ 0.909 to 0.969) and 97.4% (95% CI ‫؍‬ 0.909 to 0.969), respectively. In Burkina Faso, the sensitivity was 94.8% (95% CI ‫؍‬ 0.88.7 to 97.9%), and the specificity was 82.4% (95% CI ‫؍‬ 75.4 to 87.7%) compared to microscopy and 93.3% (95% CI ‫؍‬ 87.4 to 96.7%) and 91.4% (95% CI ‫؍‬ 85.2 to 95.3%) compared to RDT. In Thailand, the sensitivity and specificity were 93.4% (CI ‫؍‬ 86.4 to 97.1%) and 90.9 (95% CI ‫؍‬ 86.7 to 93.9%), respectively, compared to microscopy and 95.6% (95% CI ‫؍‬ 88.5 to 98.6%) and 87.1% (95% CI ‫؍‬ 82.5 to 90.6) compared to RDT. db-PCR-NALFIA is highly sensitive and specific for easy and rapid detection of Plasmodium parasites and can be easily used in countries where malaria is endemic. The inability of the device to discriminate Plasmodium species requires further investigation.

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Real-time PCR assay and rapid diagnostic tests for the diagnosis of clinically suspected malaria patients in Bangladesh

Cited by 59 publications

References 28 publications

Mutations in Plasmodium falciparum K13 propeller gene from Bangladesh (2009–2013)

Mutations in Plasmodium falciparum K13 propeller gene from Bangladesh (2009–2013)

Performance of a HRP-2/pLDH based rapid diagnostic test at the Bangladesh-India-Myanmar border areas for diagnosis of clinical malaria

Direct Blood PCR in Combination with Nucleic Acid Lateral Flow Immunoassay for Detection of Plasmodium Species in Settings Where Malaria Is Endemic

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