Characterization of Plasmodium Lactate Dehydrogenase and Histidine-Rich Protein 2 Clearance Patterns via Rapid On-Bead Detection from a Single Dried Blood Spot

Markwalter, Christine F.; Gibson, Lauren E.; Mudenda, Lwiindi; Kimmel, Danielle W.; Mbambara, Saidon; Thuma, Philip E.; Wright, David W.

doi:10.4269/ajtmh.17-0996

Cited by 23 publications

(26 citation statements)

References 33 publications

(43 reference statements)

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“…The concentration of eluted PfHRP2 from DBS to be equivalent to approximately a 1:20 dilution from whole blood, similarly to previously reported data (40). Differently, for pLDH we found that antigen concentration in eluted DBS corresponds to a 1:60 whole blood dilution, which differs from previously published data showing no differences in antigen recovery between PfHRP2 and pLDH (20). However, such differences could be explained by the different extraction methodologies and storage conditions used.…”

Section: Discussioncontrasting

confidence: 99%

“…Additionally, the assay can also quantify Pv pLDH down to 1211.6 pg/ml. The assay analytical sensitivity to detect PfHRP2 is comparable to that of a recently developed bead suspension assay based on Luminex technology (25), as well as to other immunoassays that use different technologies (20,28). This suggests that with the current technology available for the quantification of PfHRP2 using antibodies, the lowest limit of detection achievable is in the range of 0.5 to 10 pg/ml.…”

Section: Discussionmentioning

confidence: 79%

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Quantification of malaria antigens PfHRP2 and pLDH by quantitative suspension array technology in whole blood, dried blood spot and plasma

Martiáñez–Vendrell

Jiménez

Vásquez

et al. 2019

Preprint

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BackgroundMalaria diagnostics by rapid diagnostic tests (RDTs) relies primarily on the qualitative detection of Plasmodium falciparum histidine-rich protein 2 (PfHRP2) and Plasmodium sp lactate dehydrogenase (pLDH). As novel RDTs with increased sensitivity are being developed and implemented as point of care diagnostics, highly sensitive laboratory based assays are needed for evaluating RDTs performance. Here, a quantitative suspension array technology (qSAT) was developed, validated and applied for the simultaneous detection of PfHRP2 and pLDH in a variety of clinical samples (whole blood, plasma and dried blood spots) from different endemic countries.ResultsThe qSAT was specific for the target antigens, with analytical ranges of 6.8 to 762.8 pg/ml for PfHRP2 and 78.1 to 17076.6 pg/ml for P. falciparum (Pf-LDH). The assay detected P. vivax LDH (Pv-LDH) at a lower sensitivity than Pf-LDH (analytical range of 1093.20 to 187288.5 pg/ml). Both PfHRP2 and pLDH levels determined using the qSAT showed to positively correlate with parasite densities determined by quantitative PCR (Spearman r=0.59 and 0.75, respectively) as well as microscopy (Spearman r=0.40 and 0.75, respectively), suggesting the assay to be a good predictor of parasite density.ConclusionThis immunoassay can be used as a reference test for the detection and quantification of PfHRP2 and pLDH, and could serve for external validation of RDTs performance, to determine antigen persistence after parasite clearance, as well as a complementary tool to assess malaria burden in endemic settings.

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Section: Discussioncontrasting

confidence: 99%

Section: Discussionmentioning

confidence: 79%

Quantification of malaria antigens PfHRP2 and pLDH by quantitative suspension array technology in whole blood, dried blood spot and plasma

Martiáñez–Vendrell

Jiménez

Vásquez

et al. 2019

Preprint

View full text Add to dashboard Cite

show abstract

“…Additionally, the assay can also quantify P. vivax pLDH down to 1211.6 pg/ml. The assay analytical sensitivity to detect PfHRP2 is comparable to that of a recently developed bead suspension assay based on Luminex technology [25], as well as to other immunoassays that use different technologies [20,27]. This suggests that with the current technology available for the quantification of PfHRP2 using antibodies, the lowest limit of detection achievable is in the range of 0.5-10 pg/ ml.…”

Section: Discussionmentioning

confidence: 68%

“…In contrast to PfHRP2, pLDH does not persist in blood after clearance of malaria infections and is therefore a better marker of acute and current infection [19]. Upon treatment, pLDH clearance in blood has been shown to closely track with that of parasites, suggesting pLDH to be a suitable predictor for treatment failure [20]. However, sensitivity of RDTs based on this antigen is generally lower than that of PfHRP2-based RDTs [21].…”

mentioning

confidence: 99%

Quantification of malaria antigens PfHRP2 and pLDH by quantitative suspension array technology in whole blood, dried blood spot and plasma

Martiáñez–Vendrell

Jiménez

Vásquez

et al. 2020

Malar J

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Background: Malaria diagnostics by rapid diagnostic test (RDT) relies primarily on the qualitative detection of Plasmodium falciparum histidine-rich protein 2 (PfHRP2) and Plasmodium spp lactate dehydrogenase (pLDH). As novel RDTs with increased sensitivity are being developed and implemented as point of care diagnostics, highly sensitive laboratory-based assays are needed for evaluating RDT performance. Here, a quantitative suspension array technology (qSAT) was developed, validated and applied for the simultaneous detection of PfHRP2 and pLDH in a variety of biological samples (whole blood, plasma and dried blood spots) from individuals living in different endemic countries. Results: The qSAT was specific for the target antigens, with analytical ranges of 6.8 to 762.8 pg/ml for PfHRP2 and 78.1 to 17076.6 pg/ml for P. falciparum LDH (Pf-LDH). The assay detected Plasmodium vivax LDH (Pv-LDH) at a lower sensitivity than Pf-LDH (analytical range of 1093.20 to 187288.5 pg/ml). Both PfHRP2 and pLDH levels determined using the qSAT showed to positively correlate with parasite densities determined by quantitative PCR (Spearman r = 0.59 and 0.75, respectively) as well as microscopy (Spearman r = 0.40 and 0.75, respectively), suggesting the assay to be a good predictor of parasite density. Conclusion: This immunoassay can be used as a reference test for the detection and quantification of PfHRP2 and pLDH, and could serve for external validation of RDT performance, to determine antigen persistence after parasite clearance, as well as a complementary tool to assess malaria burden in endemic settings.

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“…Because PfHRP2 antigenemia can persist for weeks, they can produce false-positive results well after resolution of infection. (6) Additionally, increasing reports of false-negative RDT results due to P. falciparum parasites that escape detection due to deletion mutations of the pfhrp2 and pfhrp3 genes raise concern that PfHRP2-based RDTs may be threatened in select regions of Africa. (7, 8) While microscopy is the traditional gold-standard for malaria diagnosis, it is only sporadically available throughout much of Africa because performance is highly operator dependent, it is labor-intensive, and it requires careful, sustained training of personnel.…”

Section: Introductionmentioning

confidence: 99%

A novel CRISPR-based malaria diagnostic capable ofPlasmodiumdetection, speciation, and drug-resistance genotyping

Hennelly

et al. 2020

Preprint

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One-sentence summary: Novel malaria SHERLOCK assays enabled robust detection, 16 speciation, and genotyping of Plasmodium spp. in diverse samples collected in Africa Abstract word count: 228 20 Manuscript word count: 5291 21 # Corresponding author information: Jonathan B. Parr, MD, MPH; 130 Mason Farm 22Rd., Chapel HillABSTRACT 24 25 CRISPR-based diagnostics are a new class of highly sensitive and specific assays with 26 multiple applications in infectious disease diagnosis. SHERLOCK, Sensitivity Enzymatic Reporter UnLOCKing, is one such CRISPR-based diagnostic that 28 combines recombinase polymerase pre-amplification, CRISPR-RNA base-pairing, and 29LwCas13a activity for nucleic acid detection. We developed SHERLOCK assays for 30 malaria capable of detecting all Plasmodium species known to cause malaria in humans 31 and species-specific detection of P. vivax and P. falciparum, the species responsible for 32 the majority of malaria cases worldwide. We validated these assays against parasite 33 genomic DNA and achieved analytical sensitivities ranging from 2.5-18.8 parasites per 34 reaction. We further tested these assays using a diverse panel of 123 clinical samples 35 from the Democratic Republic of the Congo, Uganda, and Thailand and pools of 36 Anopheles mosquitoes from Thailand. When compared to real-time PCR, the P. 37 falciparum assay achieved 94% sensitivity and 94% specificity in clinical samples. In 38 addition, we developed a SHERLOCK assay capable of detecting the dihydropteroate 39 synthetase (dhps) single nucleotide variant A581G associated with P. falciparum 40 sulfadoxine-pyrimethamine resistance. Compared to amplicon-based deep sequencing, 41 the dhps SHERLOCK assay achieved 73% sensitivity and 100% specificity when 42 applied to a panel of 43 clinical samples, with false-negative calls only at lower parasite 43 densities. These novel SHERLOCK assays have potential to spawn a new generation of 44 molecular diagnostics for malaria and demonstrate the versatility of CRISPR-based 45 diagnostic approaches. 46 47 48 Newly developed technologies that utilize Clustered, Regularly-Interspaced Palindromic 49 Repeat (CRISPR) systems have the potential to revolutionize infectious disease 50 diagnostic testing.(1) SHERLOCK, or Specific High-Sensitivity Enzymatic Reporter 51UnLOCKing, is a CRISPR-based diagnostic assay that has now been used to detect 52 dengue and Zika viruses with excellent sensitivity and specificity.(2) Its simple workflow 53 and robust performance characteristics have enabled multiplexed detection and 54 genotyping of Zika and dengue viruses, with increasingly streamlined protocols that 55 facilitate use at the point-of-care.(3, 4) SHERLOCK's potential is perhaps greatest in 56 low-resource settings where improved, reliable diagnostics are urgently needed for 57 multiple pathogens and for malaria, in particular. 58 59 Timely and accurate diagnosis is an important component of malaria control and 60 elimination efforts. The current generation of rapid diagnostic tests (RDTs) that detect 61 ...

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Characterization of Plasmodium Lactate Dehydrogenase and Histidine-Rich Protein 2 Clearance Patterns via Rapid On-Bead Detection from a Single Dried Blood Spot

Cited by 23 publications

References 33 publications

Quantification of malaria antigens PfHRP2 and pLDH by quantitative suspension array technology in whole blood, dried blood spot and plasma

Quantification of malaria antigens PfHRP2 and pLDH by quantitative suspension array technology in whole blood, dried blood spot and plasma

Quantification of malaria antigens PfHRP2 and pLDH by quantitative suspension array technology in whole blood, dried blood spot and plasma

A novel CRISPR-based malaria diagnostic capable ofPlasmodiumdetection, speciation, and drug-resistance genotyping

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