Rapid Diagnostic Tests Brands Research Articles

Optimisation and evaluation of viral genomic sequencing of SARS-CoV-2 rapid diagnostic tests: a laboratory and cohort-based study

Sequencing of SARS-CoV-2 from rapid diagnostic tests (RDTs) can bolster viral genomic surveillance efforts; however, approaches to maximise and standardise pathogen genome recovery from RDTs remain underdeveloped. We aimed to systematically optimise the elution of genetic material from RDT components and to evaluate the efficacy of RDT sequencing for outbreak investigation. In this laboratory and cohort-based study we seeded RDTs with inactivated SARS-CoV-2 to optimise the elution of genomic material from RDT lateral flow strips. We measured the effect of changes in buffer type, time in buffer, and rotation on PCR cycle threshold (Ct) value. We recruited individuals older than 18 years residing in the greater Boston area, MA, USA, from July 18 to Nov 5, 2022, via email advertising to students and staff at Harvard University, MA, USA,and via broad social media advertising. All individuals recruited were within 5days of a positive diagnostic test for SARS-CoV-2; no other relevant exclusion criteria were applied. Each individual completed two RDTs and one PCR swab. On Dec 29, 2022, we also collected RDTs from a convenience sample of individuals who were positive for SARS-CoV-2 and associated with an outbreak at a senior housing facility in MA, USA. We extracted all returned PCR swabs and RDT components (ie, swab, strip, or buffer); samples with a Ct of less than 40 were subject to amplicon sequencing. We compared the efficacy of elution and sequencing across RDT brands and components and used RDT-derived sequences to infer transmission links within the outbreak at the senior housing facility. We conducted metagenomic sequencing of negative RDTs from symptomatic individuals living in the senior housing facility. Neither elution duration of greater than 10 min nor rotation during elution impacted viral titres. Elution in Buffer AVL (Ct=31·4) and Tris-EDTA Buffer (Ct=30·8) were equivalent (p=0·34); AVL outperformed elution in lysis buffer and 50% lysis buffer (Ct=40·0, p=0·0029 for both) as well as Universal Viral Transport Medium (Ct=36·7, p=0·079). Performance of RDT strips was poorer than that of matched PCR swabs (mean Ct difference 10·2 [SD 4·3], p<0·0001); however, RDT swabs performed similarly to PCR swabs (mean Ct difference 4·1 [5·2], p=0·055). No RDT brand significantly outperformed another. Across sample types, viral load predicted the viral genome assembly length. We assembled greater than 80% complete genomes from 12 of 17 RDT-derived swabs, three of 18 strips, and four of 11 residual buffers. We generated outbreak-associated SARS-CoV-2 genomes using both amplicon and metagenomic sequencing and identified multiple introductions of the virus that resulted in downstream transmission. RDT-derived swabs are a reasonable alternative to PCR swabs for viral genomic surveillance and outbreakinvestigation. RDT-derived lateral flow strips yield accurate, but significantly fewer, viral reads than matched PCR swabs. Metagenomic sequencing of negative RDTs can identify viruses that might underlie patient symptoms. The National Science Foundation, the Hertz Foundation, the National Institute of General Medical Sciences, Harvard Medical School, the Howard Hughes Medical Institute, the US Centers for Disease Control and Prevention, the Broad Institute and the National Institute of Allergy and Infectious Diseases.

Lesson Learned from Mass Antibody Rapid Diagnostic Used in the Early COVID-19 Pandemic in Indonesia Contributors.

Laboratory examination is extremely important in handling the COVID-19 pandemic. In the first era of the pandemic, the molecular and antigen tests were limited. Hence, at that time, it was necessary to carry out antibody Rapid Diagnostic Tests (RDT). However, many antibody RDTs were yet to obtain Food and Drug Authorization (FDA)'s approval. Therefore, The Indonesian Association of Clinical Pathology and Medical Laboratory (PDS PatKLIn) decided to conduct a validity test of RDT antibodies to find out the quality of SARS-CoV-2 diagnosis performance based on these RDTs used. This is a descriptive observational design with diagnostic analysis. The retrospective secondary data were collected from 34 provinces in Indonesia from May to June 2020. Data analysis was carried out on the sensitivity and specificity values of each antibody RDT brand to the RT-PCR result and analyzed descriptive data. The amount of secondary data of antibody RDT and RT-PCR results collected was 139,908, consisting of 59 RDT brands of which 44% were authorized by The Indonesian COVID-19 Response Acceleration Task Force (Gugus Tugas Percepatan Penanganan COVID-19 Indonesia). There were huge variations of SARS-CoV-2 antibody RDT performance between total antibody types (sensitivity 59.18%, specificity 62%), IgM RDT (sensitivity 16-100%, specificity 7-97%), and RDT IgG (sensitivity 33-96%, specificity 19-100%). The variations in the RDT antibodies'performance can cause errors in diagnosis leading to significant material and immaterial losses. Therefore, cooperation from various parties is needed for the pre- and post-marketing surveillance process to assess the performance and the characteristics of each RDT kit and other diagnostic methods to assist the rapid pandemic response process.

Poor performance of malaria rapid diagnostic tests for the detection of Plasmodium malariae among returned international travellers in China

BackgroundMalaria is a worldwide infectious disease. For countries that have achieved malaria elimination, the prevention of re-establishment due to infections in returned travellers has become important. The accurate and timely diagnosis of malaria is the key in preventing re-establishment, and malaria rapid diagnostic tests (RDTs) are frequently used due to their convenience. However, the RDT performance in Plasmodium malariae (P. malariae) infection diagnosis remains unknown.MethodsThis study analysed epidemiological features and diagnosis patterns of imported P. malariae cases from 2013 to 2020 in Jiangsu Province and evaluated the sensitivity of four parasite enzyme lactate dehydrogenase (pLDH)-targeting RDTs (Wondfo, SD BIONLINE, CareStart and BioPerfectus) and one aldolase-targeting RDT(BinaxNOW) for P. malariae detection. Furthermore, influential factors were investigated, including parasitaemia load, pLDH concentration and target gene polymorphisms.ResultsThe median duration from symptom onset to diagnosis among patients with P. malariae infection was 3 days, which was longer than that with Plasmodium falciparum (P. falciparum) infection. The RDTs had a low detection rate (39/69, 56.5%) among P. malariae cases. All tested RDT brands had poor performance in P. malariae detection. All the brands except the worst-performing SD BIOLINE, achieved 75% sensitivity only when the parasite density was higher than 5000 parasites/μL. Both pLDH and aldolase showed relatively conserved and low gene polymorphism rates.ConclusionsThe diagnosis of imported P. malariae cases was delayed. The RDTs had poor performance in P. malariae diagnosis and may threaten the prevention of malaria re-establishment from returned travellers. The improved RDTs or nucleic acid tests for P. malariae cases are urgently needed for the detection of imported cases in the future.

Performance of rapid rk39 tests for the diagnosis of visceral leishmaniasis in Ethiopia: a systematic review and meta-analysis

BackgroundVisceral Leishmaniasis (VL) is a severely neglected disease affecting millions of people with high mortality if left untreated. In Ethiopia, the primary laboratory diagnosis of VL is by using an antigen from a 39-amino acid sequence repeat of a kinesin-related (rK39) of leishmania donovani complex (L. donovani), rapid diagnostic tests (RDT). Different rk39 RDT brands are available with very variable performance and studies from Ethiopia showed a very wide range of sensitivity and specificity. Therefore, a systematic review and meta-analysis were conducted to determine the pooled sensitivity and specificity of rk39 RDT in Ethiopia.MethodPUBMED, EMBASE, and other sources were searched using predefined search terms to retrieve all relevant articles from 2007 to 2020. Heterogeneity was assessed by visually inspecting summary receiver operating curves (SROC), Spearman correlation coefficient (rs), Cochran Q test statistics, inconsistency square (I2) and subgroup analysis. The presence and statistical significance of publication bias were assessed by Egger's test at p < 0.05, and all the measurements showed the presence of considerable heterogeneity. Quality assessment of diagnostic accuracy studies (QUADAS-2) checklists was used to check the qualities of the study.ResultsA total of 664 articles were retrieved, and of this 12 articles were included in the meta-analysis. Overall pooled sensitivity and specificity of the rk39 RDT to diagnose VL in Ethiopia were 88.0% (95% CI 86.0% to 89.0%) and 84.0% (95% CI 82.0% to 86.0%), respectively. The sensitivity and specificity of the rk39 RDT commercial test kits were DiaMed: 86.9% (95% CI 84.3% to 89.1%) and 82.2% (95% CI 79.3% to 85.0%), and InBios: 80.0% (95% CI 77.0% to 82.8%) and 97.4% (95% CI 95.0% to 98.8%), respectively.ConclusionReferring to our result, rk39 RDT considered an essential rapid diagnostic test for VL diagnosis. Besides to the diagnostic accuracy, the features such as easy to perform, quick (10–20 min), cheap, equipment-free, electric and cold chain free, and result reproducibility, rk39 RDT is advisable to remains in practice as a diagnostic test at least in the remote VL endemic localities till a better test will come.

Malaria Rapid Diagnostic Tests: Literary Review and Recommendation for a Quality Assurance, Quality Control Algorithm.

Malaria rapid diagnostic tests (RDTs) have had an enormous global impact which contributed to the World Health Organization paradigm shift from empiric treatment to obtaining a parasitological diagnosis prior to treatment. Microscopy, the classic standard, requires significant expertise, equipment, electricity, and reagents. Alternatively, RDT’s lower complexity allows utilization in austere environments while achieving similar sensitivities and specificities. Worldwide, there are over 200 different RDT brands that utilize three antigens: Plasmodium histidine-rich protein 2 (PfHRP-2), Plasmodium lactate dehydrogenase (pLDH), and Plasmodium aldolase (pALDO). pfHRP-2 is produced exclusively by Plasmodium falciparum and is very Pf sensitive, but an alternative antigen or antigen combination is required for regions like Asia with significant Plasmodium vivax prevalence. RDT sensitivity also decreases with low parasitemia (<100 parasites/uL), genetic variability, and prozone effect. Thus, proper RDT selection and understanding of test limitations are essential. The Center for Disease Control recommends confirming RDT results by microscopy, but this is challenging, due to the utilization of clinical laboratory standards, like the College of American Pathologists (CAP) and the Clinical Lab Improvement Act (CLIA), and limited recourses. Our focus is to provide quality assurance and quality control strategies for resource-constrained environments and provide education on RDT limitations.

Rapid diagnostic tests for Plasmodium vivax malaria in endemic countries.

Rapid diagnostic tests for Plasmodium vivax malaria in endemic countries.

False-negative malaria rapid diagnostic test results and their impact on community-based malaria surveys in sub-Saharan Africa

Surveillance and diagnosis of Plasmodium falciparum malaria relies predominantly on rapid diagnostic tests (RDT). However, false-negative (FN) RDT results are known to occur for a variety of reasons, including operator...

Comparison of four rapid diagnostic tests, ELISA, microscopy and PCR for the detection of Giardia lamblia, Cryptosporidium spp. and Entamoeba histolytica in feces

PurposeMicroscopy is the diagnostic reference standard for the detection of parasites, but it is labor-intensive and requires experience. Rapid diagnostic tests (RDTs) can provide an alternative to microscopy. MethodsRDTs from four different manufacturers were compared to enzyme-linked immunosorbent assay (ELISA), microscopy and/or parasite-specific real-time PCR: ImmunoCardSTAT!®CGE (Meridian Bioscience Inc., Cincinnati, Ohio, USA) (A), Crypto/Giardia Duo-Strip (Coris Bioconcepts, Gembloux, Belgium) (B), RIDA®QUICK Cryptosporidium/Giardia/Entamoeba Combi (R-BioPharm, Darmstadt, Germany) (C) and Giardia/Cryptosporidium Quik Chek (Techlab Inc., Blacksburg, Virginia, USA) (D). ResultsThirty frozen samples were analyzed retrospectively. For Giardia lamblia (n=12) and Cryptosporidium (n=12) sensitivities ranged from 58% (B), over 83% (A, C) to 100% (D) and from 92% (B) to 100% (A, C, D), respectively. Specificity for both G. lamblia and Cryptosporidium was 100% for all RDT brands. Sensitivity for Entamoeba histolytica (n=5) was 100%, while specificity reached 80% (A) to 88% (C). In a prospective study, fresh samples were tested. For G. lamblia (n=30), sensitivity ranged from 66% (B), over 79% (A) and 83% (C) to 100% (D) and specificity varied between 94% (D) and 100% (A, B, C). For Cryptosporidium (n=3), sensitivity was 100% for all brands except (B) (67%) and specificities were 95% (A, B), 98% (C) and 100% (D). E. histolytica (n=1) was detected by both (A) and (C), while specificity was 81% and 87% respectively. ConclusionRDTs can be a valuable tool when microscopic expertise is poor and in remote and outbreak settings where other techniques are often not available and rapid diagnosis is required.

Lignes directrices de l’association canadienne des pathologistes concernant l’implantation d’un service de télépathologie diagnostique à l’aide de lames virtuelles

Microscopy is the diagnostic reference standard for the detection of parasites, but it is labor-intensive and requires experience. Rapid diagnostic tests (RDTs) can provide an alternative to microscopy.RDTs from four different manufacturers were compared to enzyme-linked immunosorbent assay (ELISA), microscopy and/or parasite-specific real-time PCR: ImmunoCardSTAT!®CGE (Meridian Bioscience Inc., Cincinnati, Ohio, USA) (A), Crypto/Giardia Duo-Strip (Coris Bioconcepts, Gembloux, Belgium) (B), RIDA®QUICK Cryptosporidium/Giardia/Entamoeba Combi (R-BioPharm, Darmstadt, Germany) (C) and Giardia/Cryptosporidium Quik Chek (Techlab Inc., Blacksburg, Virginia, USA) (D).Thirty frozen samples were analyzed retrospectively. For Giardia lamblia (n = 12) and Cryptosporidium (n = 12) sensitivities ranged from 58% (B), over 83% (A, C) to 100% (D) and from 92% (B) to 100% (A, C, D), respectively. Specificity for both G. lamblia and Cryptosporidium was 100% for all RDT brands. Sensitivity for Entamoeba histolytica (n = 5) was 100%, while specificity reached 80% (A) to 88% (C). In a prospective study, fresh samples were tested. For G. lamblia (n = 30), sensitivity ranged from 66% (B), over 79% (A) and 83% (C) to 100% (D) and specificity varied between 94% (D) and 100% (A, B, C). For Cryptosporidium (n = 3), sensitivity was 100% for all brands except (B) (67%) and specificities were 95% (A, B), 98% (C) and 100% (D). E. histolytica (n = 1) was detected by both (A) and (C), while specificity was 81% and 87% respectively.RDTs can be a valuable tool when microscopic expertise is poor and in remote and outbreak settings where other techniques are often not available and rapid diagnosis is required.

Use of Rapid Diagnostic Tests in Malaria School Surveys in Kenya: Does their Under-performance Matter for Planning Malaria Control?

Malaria rapid diagnostic tests (RDTs) are known to yield false-positive results, and their use in epidemiologic surveys will overestimate infection prevalence and potentially hinder efficient targeting of interventions. To examine the consequences of using RDTs in school surveys, we compared three RDT brands used during a nationwide school survey in Kenya with expert microscopy and investigated the cost implications of using alternative diagnostic approaches in identifying localities with differing levels of infection. Overall, RDT sensitivity was 96.1% and specificity was 70.8%. In terms of classifying districts and schools according to prevalence categories, RDTs were most reliable for the < 1% and > 40% categories and least reliable in the 1–4.9% category. In low-prevalence settings, microscopy was the most expensive approach, and RDT results corrected by either microscopy or polymerase chain reaction were the cheapest. Use of polymerase chain reaction–corrected RDT results is recommended in school malaria surveys, especially in settings with low-to-moderate malaria transmission.

Prozone in malaria rapid diagnostics tests: how many cases are missed?

BackgroundProzone means false-negative or false-low results in antigen-antibody reactions, due to an excess of either antigen or antibody. The present study prospectively assessed its frequency for malaria rapid diagnostic tests (RDTs) and Plasmodium falciparum samples in an endemic field setting.MethodsFrom January to April 2010, blood samples with P. falciparum high parasitaemia (≥ 4% red blood cells infected) were obtained from patients presenting at the Provincial Hospital of Tete (Mozambique). Samples were tested undiluted and 10-fold diluted in saline with a panel of RDTs and results were scored for line intensity (no line visible, faint, weak, medium and strong). Prozone was defined as a sample which showed no visible test line or a faint or weak test line when tested undiluted, and a visible test line of higher intensity when tested 10-fold diluted, as observed by two blinded observers and upon duplicate testing.ResultsA total of 873/7,543 (11.6%) samples showed P. falciparum, 92 (10.5%) had high parasitaemia and 76 were available for prozone testing. None of the two Pf-pLDH RDTs, but all six HRP-2 RDTs showed prozone, at frequencies between 6.7% and 38.2%. Negative and faint HRP-2 lines accounted for four (3.8%) and 15 (14.4%) of the 104 prozone results in two RDT brands. For the most affected brand, the proportions of prozone with no visible or faint HRP-2 lines were 10.9% (CI: 5.34-19.08), 1.2% (CI: 0.55-2.10) and 0.1% (CI: 0.06-0.24) among samples with high parasitaemia, all positive samples and all submitted samples respectively. Prozone occurred mainly, but not exclusively, among young children.ConclusionProzone occurs at different frequency and intensity in HRP-2 RDTs and may decrease diagnostic accuracy in the most affected RDTs.

Rapid diagnostic tests as a source of DNA for Plasmodium species-specific real-time PCR

BackgroundThis study describes the use of malaria rapid diagnostic tests (RDTs) as a source of DNA for Plasmodium species-specific real-time PCR.MethodsFirst, the best method to recover DNA from RDTs was investigated and then the applicability of this DNA extraction method was assessed on 12 different RDT brands. Finally, two RDT brands (OptiMAL Rapid Malaria Test and SDFK60 malaria Ag Plasmodium falciparum/Pan test) were comprehensively evaluated on a panel of clinical samples submitted for routine malaria diagnosis at ITM. DNA amplification was done with the 18S rRNA real-time PCR targeting the four Plasmodium species. Results of PCR on RDT were compared to those obtained by PCR on whole blood samples.ResultsBest results were obtained by isolating DNA from the proximal part of the nitrocellulose component of the RDT strip with a simple DNA elution method. The PCR on RDT showed a detection limit of 0.02 asexual parasites/μl, which was identical to the same PCR on whole blood. For all 12 RDT brands tested, DNA was detected except for one brand when a low parasite density sample was applied. In RDTs with a plastic seal covering the nitrocellulose strip, DNA extraction was hampered. PCR analysis on clinical RDT samples demonstrated correct identification for single species infections for all RDT samples with asexual parasites of P. falciparum (n = 60), Plasmodium vivax (n = 10), Plasmodium ovale (n = 10) and Plasmodium malariae (n = 10). Samples with only gametocytes were detected in all OptiMAL and in 10 of the 11 SDFK60 tests. None of the negative samples (n = 20) gave a signal by PCR on RDT. With PCR on RDT, higher Ct-values were observed than with PCR on whole blood, with a mean difference of 2.68 for OptiMAL and 3.53 for SDFK60. Mixed infections were correctly identified with PCR on RDT in 4/5 OptiMAL tests and 2/5 SDFK60 tests.ConclusionsRDTs are a reliable source of DNA for Plasmodium real-time PCR. This study demonstrates the best method of RDT fragment sampling for a wide range of RDT brands in combination with a simple and low cost extraction method, allowing RDT quality control.

External quality assessment on the use of malaria rapid diagnostic tests in a non-endemic setting

BackgroundMalaria rapid diagnostic tests (RDTs) are increasingly used as a tool for the diagnosis of malaria, both in endemic and in non-endemic settings. The present study reports the results of an external quality assessment (EQA) session on RDTs in a non-endemic setting.MethodsAfter validation of antigen stability during shipment at room temperature, three clinical samples and a questionnaire were sent to clinical laboratories in Belgium and the Grand Duchy of Luxembourg using malaria RDTs. Participants were asked to report the results of the RDTs as observations (visibility of the RDT control and test lines) and interpretations (report as formulated to the clinician). In addition, participants were invited to fill in a questionnaire on the place of RDTs in the diagnostic strategy of malaria.ResultsA total of 128/133 (96.2%) of clinical laboratories using RDTs participated. Six three-band and one four-band RDT brands were used. Analytical errors were rare and included (i) not recognizing invalid RDT results (1.6%) and (ii) missing the diagnosis of Plasmodium falciparum (0.8%). Minor errors were related to RDT test result interpretation and included (i) reporting "RDT positive" without species identification in the case of P. falciparum and non-falciparum species (16.9% and 6.5% respectively) and (ii) adding incorrect comments to the report (3.2%). Some of these errors were related to incorrect RDT package insert instructions such as (i) not reporting the possibility of mixed species infection in the case of P. falciparum and Plasmodium vivax (35.5% and 18.5% respectively) and (ii) the interpretation of P. vivax instead of non-falciparum species at the presence of a pan-species antigen line (4.0%). According to the questionnaire, 48.8% of participants processed ≤20 requests for malaria diagnosis in 2009. During opening hours, 93.6% of 125 participants used RDTs as an adjunct to microscopy but outside opening hours, nearly one third of 113 participants relied on RDTs as the primary (4.4%) or the single tool (25.7%) for malaria diagnosis.ConclusionIn this non-endemic setting, errors in RDT performance were mainly related to RDT test line interpretations, partly due to incorrect package insert instructions. The reliance on RDTs as the primary or the single tool for the diagnosis of malaria outside opening hours is of concern and should be avoided.

Buffer substitution in malaria rapid diagnostic tests causes false-positive results

BackgroundMalaria rapid diagnostic tests (RDTs) are kits that generally include 20 to 25 test strips or cassettes, but only a single buffer vial. In field settings, laboratory staff occasionally uses saline, distilled water (liquids for parenteral drugs dilution) or tap water as substitutes for the RDT kit's buffer to compensate for the loss of a diluent bottle. The present study assessed the effect of buffer substitution on the RDT results.MethodsTwenty-seven RDT brands were run with EDTA-blood samples of five malaria-free subjects, who were negative for rheumatoid factor and antinuclear antibodies. Saline, distilled water and tap water were used as substitute liquids. RDTs were also run with distilled water, without adding blood. Results were compared to those obtained with the RDT kit's buffer and Plasmodium positive samples.ResultsOnly eight cassettes (in four RDT brands) showed no control line and were considered invalid. Visible test lines occurred for at least one malaria-free sample and one of the substitutes in 20/27 (74%) RDT brands (saline: n = 16; distilled water: n = 17; and tap water: n = 20), and in 15 RDTs which were run with distilled water only. They occurred for all Plasmodium antigens and RDT formats (two-, three- and four-band RDTs). Clearance of the background of the strip was excellent except for saline. The aspects (colour, intensity and crispness) of the control and the false-positive test lines were similar to those obtained with the RDT kits' buffer and Plasmodium positive samples.ConclusionReplacement of the RDT kit's dedicated buffer by saline, distilled water and tap water can cause false-positive test results.

Malaria rapid diagnostic tests: Plasmodium falciparum infections with high parasite densities may generate false positive Plasmodium vivax pLDH lines

BackgroundMost malaria rapid diagnostic tests (RDTs) detect Plasmodium falciparum and an antigen common to the four species. Plasmodium vivax-specific RDTs target P. vivax-specific parasite lactate dehydrogenase (Pv-pLDH). Previous observations of false positive Pv-pLDH test lines in P. falciparum samples incited to the present study, which assessed P. vivax-specific RDTs for the occurrence of false positive Pv-pLDH lines in P. falciparum samples.MethodsNine P. vivax-specific RDTs were tested with 85 P. falciparum samples of high (≥2%) parasite density. Mixed P. falciparum/P. vivax infections were ruled out by real-time PCR. The RDTs included two-band (detecting Pv-pLDH), three-band (detecting P. falciparum-antigen and Pv-pLDH) and four-band RDTs (detecting P. falciparum, Pv-pLDH and pan-pLDH).ResultsFalse positive Pv-pLDH lines were observed in 6/9 RDTs (including two- three- and four-band RDTs). They occurred in the individual RDT brands at frequencies ranging from 8.2% to 29.1%. For 19/85 samples, at least two RDT brands generated a false positive Pv-pLDH line. Sixteen of 85 (18.8%) false positive lines were of medium or strong line intensity. There was no significant relation between false positive results and parasite density or geographic origin of the samples.ConclusionFalse positive Pv-pLDH lines in P. falciparum samples with high parasite density occurred in 6/9 P. vivax-specific RDTs. This is of concern as P. falciparum and P. vivax are co-circulating in many regions. The diagnosis of life-threatening P. falciparum malaria may be missed (two-band Pv-pLDH RDT), or the patient may be treated incorrectly with primaquine (three- or four-band RDTs).

Assessment of the prozone effect in malaria rapid diagnostic tests

BackgroundThe prozone effect (or high doses-hook phenomenon) consists of false-negative or false-low results in immunological tests, due to an excess of either antigens or antibodies. Although frequently cited as a cause of false-negative results in malaria rapid diagnostic tests (RDTs), especially at high parasite densities of Plasmodium falciparum, it has been poorly documented. In this study, a panel of malaria RDTs was challenged with clinical samples with P. falciparum hyperparasitaemia (> 5% infected red blood cells).MethodsTwenty-two RDT brands were tested with seven samples, both undiluted and upon 10 ×, 50 × and 100 × dilutions in NaCl 0.9%. The P. falciparum targets included histidine-rich protein-2 (HRP-2, n = 17) and P. falciparum-specific parasite lactate dehydrogenase (Pf-pLDH, n = 5). Test lines intensities were recorded in the following categories: negative, faint, weak, medium or strong. The prozone effect was defined as an increase in test line intensity of at least one category after dilution, if observed upon duplicate testing and by two readers.ResultsSixteen of the 17 HRP-2 based RDTs were affected by prozone: the prozone effect was observed in at least one RDT sample/brand combination for 16/17 HRP-2 based RDTs in 6/7 samples, but not for any of the Pf-pLDH tests. The HRP-2 line intensities of the undiluted sample/brand combinations with prozone effect (n = 51) included a single negative (1.9%) and 29 faint and weak readings (56.9%). The other target lens (P. vivax-pLDH, pan-specific pLDH and aldolase) did not show a prozone effect.ConclusionThis study confirms the prozone effect as a cause of false-negative HRP-2 RDTs in samples with hyperparasitaemia.