scholarly journals Direct Comparison of Standard and Ultrasensitive PCR for the Detection of Plasmodium falciparum from Dried Blood Spots in Bagamoyo, Tanzania

2021 ◽  
Vol 104 (4) ◽  
pp. 1371-1374
Author(s):  
Christine F. Markwalter ◽  
Billy Ngasala ◽  
Tonelia Mowatt ◽  
Christopher Basham ◽  
Zackary Park ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Diagnostic Tests ◽  
Dried Blood Spots ◽  
Rapid Diagnostic Tests ◽  
Low Density ◽  
Rt Pcr ◽  
Limited Evidence ◽  
Blood Spots ◽  
Asymptomatic Infections ◽  
Incremental Increase

ABSTRACTUltrasensitive PCR used in low-transmission malaria-endemic settings has revealed a much higher burden of asymptomatic infections than that detected by rapid diagnostic tests (RDTs) or standard PCR, but there is limited evidence as to whether this is the case in higher transmission settings. Using dried blood spots (DBS) collected among 319 schoolchildren in Bagamoyo, Tanzania, we found good correlation (Pearson’s R = 0.995) between Plasmodium falciparum parasite densities detected by a DNA-based 18s rRNA real-time PCR (qPCR) and an RNA-based ultrasensitive reverse transcriptase (RT)-PCR (usPCR) for the same target. Whereas prevalence by usPCR was higher than that found by qPCR (37% versus 32%), the proportion of additionally detected low-density infections (median parasite density < 0.050 parasites/µL) represented an incremental increase. It remains unclear to what extent these low-density infections may contribute to the infectious reservoir in different malaria transmission settings.

scholarly journals Genetic Variations of Plasmodium Falciparum Histidine-rich Protein 2 and 3 in Assosa Zone, Ethiopia: Its Impact on the Performance of Malaria Rapid Diagnostic Tests

2021 ◽  
Author(s):  
Gezahegn Solomon Alemayehu ◽  
Alebachew Messele ◽  
Karen Lopez ◽  
Eugenia Lo ◽  
Daniel Janies ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Amino Acid ◽  
Diagnostic Tests ◽  
False Negative ◽  
Dried Blood Spots ◽  
Genetic Variations ◽  
Rapid Diagnostic Tests ◽  
Repeat Type ◽  
Amino Acid Repeat ◽  
Cross Sectional

Abstract Background: Rapid diagnostic tests (RDT) are commonly used for the diagnosis of Plasmodium falciparum malaria. However, false negative results of RDT caused by genetic variations of P. falciparum histidine-rich protein 2 and 3 genes (pfhrp2/3) threaten existing malaria case management and control efforts. The main objective of this study was to investigate the genetic variations of the pfhrp2/3 genes. Methods: A cross-sectional study was conducted from malaria symptomatic individuals in 2018 in Assosa zone, Ethiopia. Finger prick samples were collected for RDT and microscopic examination of thick and thin blood films. Dried blood spots (DBS) were used for genomic parasite DNA extraction and molecular detection. Amplification of parasite DNA was made by quantitative PCR. DNA amplicons of pfhrp2/3 were purified and sequenced. Results: The PfHRP2 repeat type isolates were less conserved compared to the PfHRP3 repeat type. A total of eleven and eight different PfHRP2 and PfHRP3 amino acid repeat types were identified, respectively. Type 1, 4 and 7 repeats were shared by PfHRP2 and PfHRP3 isolates. Type 2 repeats were found only in PfHRP2, while types 16 and 17 were found only in PfHRP3 with a high frequency in all isolates. 18 novel repeat types were found in PfHRP2 and 13 novel repeat types were found in PfHRP3 in single or multiple copies per isolate. The positivity rate for PfHRP2 RDT was high, 82.9% in PfHRP2 and 84.3 % in PfHRP3 sequence isolate at parasitemia levels >250 parasites/µl. Using the Baker model, 100 % of the isolates in group A and 73.7% of the isolates in group B were predicted to be detected by PfHRP2 RDT at parasitemia level> 250 parasite/μl. Conclusion: The findings of this study indicate the presence of different PfHRP2 and PfHRP3 amino acid repeat including novel repeats in P. falciparum from Ethiopia. These results indicate that there is a need to closely monitor the performance of PfHRP2 RDT associated with the genetic variation of the pfhrp2 and pfhrp3 gene in P. falciparum isolates at the country-wide level.

scholarly journals Stratifying Malaria Receptivity In Bangladesh Using Archived Rapid Diagnostic Tests

2020 ◽  
Author(s):  
André Barembaye Sagna ◽  
Mohammad Golam Kibria ◽  
Shamsun Naher ◽  
Shayla Islam ◽  
M. M. Aktaruzzaman ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Salivary Gland ◽  
Diagnostic Tests ◽  
Rapid Diagnostic Tests ◽  
Low Density ◽  
Bed Net ◽  
Malaria Epidemiology ◽  
Net Use ◽  
Serological Studies ◽  
Epidemiology Methods

Abstract Background Surveillance on low-density infections and exposure to vectors is crucial to understand where malaria elimination might be feasible, and where the risk of outbreaks is high. Archived rapid diagnostic tests (RDTs), used by national malaria control and elimination programs for clinical diagnosis, present a valuable, yet rarely used resource for in-depth studies on malaria epidemiology. Methods We screened 1,022 RDTs from two sub-Districts in Bangladesh (Alikadam and Kamalganj) by qPCR for low-density Plasmodium falciparum and Plasmodium vivax infections, and by ELISA for Anopheles salivary gland antibodies as a marker for exposure to vectors. Results Concordance between RDT and qPCR was moderate. qPCR detected 31/1022 infections compared to 36/1022 diagnosed by RDT. Exposure to Anopheles was significantly higher in Kamalganj despite low transmission, which could be explained by low bed net use. Conclusions Archived RDTs present a valuable source of antibodies for serological studies on exposure to vectors. In contrast, the benefit of screening archived RDTs to obtain a better estimate of test positivity is moderate. Kamalganj could thus be prone to outbreaks.

scholarly journals Stratifying malaria receptivity in Bangladesh using archived rapid diagnostic tests

2020 ◽  
Vol 19 (1) ◽  
Author(s):  
André Barembaye Sagna ◽  
Mohammad Golam Kibria ◽  
Shamsun Naher ◽  
Shayla Islam ◽  
M. M. Aktaruzzaman ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Salivary Gland ◽  
Diagnostic Tests ◽  
Clinical Case ◽  
Rapid Diagnostic Tests ◽  
Low Density ◽  
Bed Net ◽  
Malaria Epidemiology ◽  
Net Use ◽  
Serological Studies

Abstract Background Surveillance of low-density infections and of exposure to vectors is crucial to understand where malaria elimination might be feasible, and where the risk of outbreaks is high. Archived rapid diagnostic tests (RDTs), used by national malaria control and elimination programs for clinical diagnosis, present a valuable, yet rarely used resource for in-depth studies on malaria epidemiology. Methods 1022 RDTs from two sub-Districts in Bangladesh (Alikadam and Kamalganj) were screened by qPCR for low-density Plasmodium falciparum and Plasmodium vivax infections, and by ELISA for Anopheles salivary gland antibodies as a marker for exposure to vectors. Results Concordance between RDT and qPCR was moderate. qPCR detected 31/1022 infections compared to 36/1022 diagnosed by RDT. Exposure to Anopheles was significantly higher in Kamalganj despite low transmission, which could be explained by low bed net use. Conclusions Archived RDTs present a valuable source of antibodies for serological studies on exposure to vectors. In contrast, the benefit of screening archived RDTs to obtain a better estimate of clinical case numbers is moderate. Kamalganj could be prone to outbreaks.

scholarly journals Antibody signatures of asymptomatic Plasmodium falciparum malaria infections measured from dried blood spots

2021 ◽  
Vol 20 (1) ◽  
Author(s):  
Christine F. Markwalter ◽  
Myat Htut Nyunt ◽  
Zay Yar Han ◽  
Ricardo Henao ◽  
Aarti Jain ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Specific Antibody ◽  
Predictive Value ◽  
Dried Blood Spots ◽  
Protein Microarrays ◽  
Antibody Responses ◽  
Antibody Activity ◽  
Low Density ◽  
Blood Spots ◽  
Diagnostic Characteristics

Abstract Background Screening malaria-specific antibody responses on protein microarrays can help identify immune factors that mediate protection against malaria infection, disease, and transmission, as well as markers of past exposure to both malaria parasites and mosquito vectors. Most malaria protein microarray work has used serum as the sample matrix, requiring prompt laboratory processing and a continuous cold chain, thus limiting applications in remote locations. Dried blood spots (DBS) pose minimal biohazard, do not require immediate laboratory processing, and are stable at room temperature for transport, making them potentially superior alternatives to serum. The goals of this study were to assess the viability of DBS as a source for antibody profiling and to use DBS to identify serological signatures of low-density Plasmodium falciparum infections in malaria-endemic regions of Myanmar. Methods Matched DBS and serum samples from a cross-sectional study in Ingapu Township, Myanmar were probed on protein microarrays populated with P. falciparum antigen fragments. Signal and trends in both sample matrices were compared. A case-control study was then performed using banked DBS samples from malaria-endemic regions of Myanmar, and a regularized logistic regression model was used to identify antibody signatures of ultrasensitive PCR-positive P. falciparum infections. Results Approximately 30% of serum IgG activity was recovered from DBS. Despite this loss of antibody activity, antigen and population trends were well-matched between the two sample matrices. Responses to 18 protein fragments were associated with the odds of asymptomatic P. falciparum infection, albeit with modest diagnostic characteristics (sensitivity 58%, specificity 85%, negative predictive value 88%, and positive predictive value 52%). Conclusions Malaria-specific antibody responses can be reliably detected, quantified, and analysed from DBS, opening the door to serological studies in populations where serum collection, transport, and storage would otherwise be impossible. While test characteristics of antibody signatures were insufficient for individual diagnosis, serological testing may be useful for identifying exposure to asymptomatic, low-density malaria infections, particularly if sero-surveillance strategies target individuals with low previous exposure as sentinels for population exposure.

scholarly journals Limitations of rapid diagnostic tests in malaria surveys in areas with varied transmission intensity in Uganda 2017-2019: Implications for selection and use of HRP2 RDTs

PLoS ONE ◽  
2020 ◽  
Vol 15 (12) ◽  
pp. e0244457
Author(s):  
Agaba B. Bosco ◽  
Joaniter I. Nankabirwa ◽  
Adoke Yeka ◽  
Sam Nsobya ◽  
Karryn Gresty ◽  
...  
Keyword(s):  
Diagnostic Tests ◽  
Gene Deletion ◽  
False Negative ◽  
Malaria Diagnosis ◽  
Dried Blood Spots ◽  
Rapid Diagnostic Tests ◽  
Low Density ◽  
Related Factors ◽  
Cross Sectional ◽  
Factors Associated

Background Plasmodium falciparum histidine-rich protein 2 (HRP2)-based rapid diagnostic tests (RDTs) are exclusively recommended for malaria diagnosis in Uganda; however, their functionality can be affected by parasite-related factors that have not been investigated in field settings. Methods Using a cross-sectional design, we analysed 219 RDT-/microscopy+ and 140 RDT+/microscopy+ dried blood spots obtained from symptomatic children aged 2–10 years from 48 districts in Uganda between 2017 and 2019. We aimed to investigate parasite-related factors contributing to false RDT results by molecular characterization of parasite isolates. ArcGIS software was used to map the geographical distribution of parasites. Statistical analysis was performed using chi-square or Fisher’s exact tests, with P ≤ 0.05 indicating significance. Odds ratios (ORs) were used to assess associations, while logistic regression was performed to explore possible factors associated with false RDT results. Results The presence of parasite DNA was confirmed in 92.5% (332/359) of the blood samples. The levels of agreement between the HRP2 RDT and PCR assay results in the (RDT+/microscopy+) and (RDT-/microscopy+) sample subsets were 97.8% (137/140) and 10.9% (24/219), respectively. Factors associated with false-negative RDT results in the (RDT-/microscopy+) samples were parasite density (<1,000/μl), pfhrp2/3 gene deletion and non-P. falciparum species (aOR 2.65, 95% CI: 1.62–4.38, P = 0.001; aOR 4.4, 95% CI 1.72–13.66, P = 0.004; and aOR 18.65, 95% CI: 5.3–38.7, P = 0.001, respectively). Overall, gene deletion and non-P. falciparum species contributed to 12.3% (24/195) and 19.0% (37/195) of false-negative RDT results, respectively. Of the false-negative RDTs results, 80.0% (156/195) were from subjects with low-density infections (< 25 parasites per 200 WBCs or <1,000/μl). Conclusion This is the first evaluation and report of the contributions of pfhrp2/3 gene deletion, non-P. falciparum species, and low-density infections to false-negative RDT results under field conditions in Uganda. In view of these findings, the use of HRP2 RDTs should be reconsidered; possibly, switching to combination RDTs that target alternative antigens, particularly in affected areas, may be beneficial. Future evaluations should consider larger and more representative surveys covering other regions of Uganda.

Performance of rapid diagnostic tests for the detection of antibodies to hepatitis C virus in whole blood collected on dried blood spots

2016 ◽  
Vol 23 (5) ◽  
pp. 399-401 ◽  
Author(s):  
L. Poiteau ◽  
A. Soulier ◽  
I. Rosa ◽  
F. Roudot-Thoraval ◽  
C. Hézode ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Whole Blood ◽  
Diagnostic Tests ◽  
Dried Blood Spots ◽  
Rapid Diagnostic Tests ◽  
Blood Spots

scholarly journals pfhrp2 and pfhrp3 Gene Deletions That Affect Malaria Rapid Diagnostic Tests for Plasmodium falciparum: Analysis of Archived Blood Samples From 3 African Countries

2019 ◽  
Vol 220 (9) ◽  
pp. 1444-1452 ◽  
Author(s):  
Rebecca Thomson ◽  
Khalid B Beshir ◽  
Jane Cunningham ◽  
Frank Baiden ◽  
Jameel Bharmal ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
18S Rdna ◽  
Diagnostic Tests ◽  
Merozoite Surface Protein ◽  
Dried Blood Spots ◽  
Rapid Diagnostic Tests ◽  
African Countries ◽  
Gene Deletions ◽  
Blood Samples ◽  
Malaria Rapid Diagnostic Tests

Abstract Background Malaria rapid diagnostic tests (mRDTs) that target histidine-rich protein 2 (HRP2) are important tools for Plasmodium falciparum diagnosis. Parasites with pfhrp2/3 gene deletions threaten the use of these mRDTs and have been reported in Africa, Asia, and South America. We studied blood samples from 3 African countries to determine if these gene deletions were present. Methods We analyzed 911 dried blood spots from Ghana (n = 165), Tanzania (n = 176), and Uganda (n = 570). Plasmodium falciparum infection was confirmed by 18S rDNA polymerase chain reaction (PCR), and pfhrp2/3 genes were genotyped. True pfhrp2/3 gene deletions were confirmed if samples were (1) microscopy positive; (2) 18S rDNA PCR positive; (3) positive for merozoite surface protein genes by PCR or positive by loop-mediated isothermal amplification; or (4) quantitative PCR positive with &gt;5 parasites/µL. Results No pfhrp2/3 deletions were detected in samples from Ghana, but deletions were identified in Tanzania (3 pfhrp2; 2 pfhrp3) and Uganda (7 pfhrp2; 2 pfhrp3). Of the 10 samples with pfhrp2 deletions, 9 tested negative by HRP2-based mRDT. Conclusions The presence of pfhrp2/3 deletions in Tanzania and Uganda, along with reports of pfhrp2/3-deleted parasites in neighboring countries, reinforces the need for systematic surveillance to monitor the reliability of mRDTs in malaria-endemic countries.

scholarly journals Genetic variation of Plasmodium falciparum histidine-rich protein 2 and 3 in Assosa zone, Ethiopia: its impact on the performance of malaria rapid diagnostic tests

2021 ◽  
Vol 20 (1) ◽  
Author(s):  
Gezahegn Solomon Alemayehu ◽  
Alebachew Messele ◽  
Kayla Blackburn ◽  
Karen Lopez ◽  
Eugenia Lo ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Genetic Variation ◽  
Amino Acid ◽  
Diagnostic Tests ◽  
False Negative ◽  
Dried Blood Spots ◽  
Rapid Diagnostic Tests ◽  
Repeat Type ◽  
Amino Acid Repeat ◽  
Cross Sectional

Abstract Background Rapid diagnostic tests (RDT) are commonly used for the diagnosis of malaria caused by Plasmodium falciparum. However, false negative results of RDT caused by genetic variation of P. falciparum histidine-rich protein 2 and 3 genes (pfhrp2/3) threaten existing malaria case management and control efforts. The main objective of this study was to investigate the genetic variations of the pfhrp2/3 genes. Methods A cross-sectional study was conducted from malaria symptomatic individuals in 2018 in Assosa zone, Ethiopia. Finger-prick blood samples were collected for RDT and microscopic examination of thick and thin blood films. Dried blood spots (DBS) were used for genomic parasite DNA extraction and molecular detection. Amplification of parasite DNA was made by quantitative PCR. DNA amplicons of pfhrp2/3 were purified and sequenced. Results The PfHRP2 amino acid repeat type isolates were less conserved compared to the PfHRP3 repeat type. Eleven and eight previously characterized PfHRP2 and PfHRP3 amino acid repeat types were identified, respectively. Type 1, 4 and 7 repeats were shared by PfHRP2 and PfHRP3 proteins. Type 2 repeats were found only in PfHRP2, while types 16 and 17 were found only in PfHRP3 with a high frequency in all isolates. 18 novel repeat types were found in PfHRP2 and 13 novel repeat types were found in PfHRP3 in single or multiple copies per isolate. The positivity rate for PfHRP2 RDT was high, 82.9% in PfHRP2 and 84.3% in PfHRP3 sequence isolates at parasitaemia levels > 250 parasites/µl. Using the Baker model, 100% of the isolates in group A (If product of types 2 × type 7 repeats ≥ 100) and 73.7% of the isolates in group B (If product of types 2 × type 7 repeats 50–99) were predicted to be detected by PfHRP2 RDT at parasitaemia level > 250 parasite/μl. Conclusion The findings of this study indicate the presence of different PfHRP2 and PfHRP3 amino acid repeat including novel repeats in P. falciparum from Ethiopia. These results indicate that there is a need to closely monitor the performance of PfHRP2 RDT associated with the genetic variation of the pfhrp2 and pfhrp3 gene in P. falciparum isolates at the country-wide level.

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