Ribosomal RNA-based diagnosis of Plasmodium falciparum malaria

We have identified useful target sites for the diagnosis of malaria infections by oligonucleotide hybridization on the small subunit RNA of Plasmodium falciparum. Acetic acid works as effectively as formaldehyde or methyl mercuric hydroxide in procedures designed to apply RNA to filters. We have confirmed the findings of others that the stability of ribosomal RNA suffices for its use as a target for diagnosis. We have achieved a detection level of at least 0.00046% parasitemia and suggest that detection of a single parasite is well within reach of this technology.     

Comparison of three antigen detection tests for diagnosis and follow-up of falciparum malaria in travellers returning to Berlin,Germany

We determined the sensitivity and specificity of three rapid immunochromatographic malarial antigen detection test systems (RDTs) for the detection of Plasmodium falciparumand assessed the quality of follow-up results. ParaSight-F and ICT Malaria detect histidine-rich protein-2 (HRP-2), whereas OptiMal detects plasmodial lactate dehydrogenase (pLDH). ParaSight-F performed with 95.1% sensitivity and 97.1% specificity (554 patients tested of whom 144 had falciparum malaria). ICT Malaria performed with 95.7% sensitivity and 99.2% specificity (718 patients tested of whom 184 had falciparum malaria). OptiMal performed with 76.2% sensitivity and 99.7% specificity (539 patients tested of whom 130 had falciparum malaria). In follow-up investigations, HRP-2 did not appear to be a useful antigen due to its long half-life, whereas pLDH offers a reasonable correlation with the presence of viable parasites in those cases initially detected. We therefore conclude that a combination of both antigens might be the best option for creating a reliable RDT for the diagnosis of falciparum malaria.     

Enzyme immunoassay for detection of antigen in acutePlasmodium falciparum malaria

A monoclonal antibody specific for an epitope of a 50 kDa Plasmodium falciparum antigen was used in an enzyme immunoassay for detection of the corresponding exo-antigen in culture supernatant and in the sera of 31 patients suffering from acute malaria. The assay was specific for Plasmodium falciparum and did not appear to be strain restricted. A parasitaemia level below 0.001 % could be detected.     

The detection and treatment of Plasmodium falciparum malaria: time for change

In most countries where malaria is endemic, P. falciparum malaria is on the rise. This is primarily due to the spread of drug-resistant strains. Drug resistance is mediated by spontaneous changes in the parasite genome that allow resistant parasites to escape the action of the drugs. The spread of drug resistance increases the transmission of malaria parasites. The consequences for the populations at risk are profound both in terms of consequences for health and economy. In order to halt the progression of drug resistance, we need to change the way antimalarials are used. As in tuberculosis and HIV/AIDS, we must use a combination of drugs for the treatment of malaria. Taking into account the pharmacokinetic and pharmacodynamic properties of the various anti-malarial agents, artemisinin-based combination therapy (ACT) seems to be the best option. This strategy should be used in conjunction with early diagnosis and appropriate vector control measures to achieve reduction in the emergence and spread of drug resistance.     

Development and evaluation of rapid urinary antigen detection tests for diagnosis of penicilliosis marneffei

Penicilliosis, caused by the dimorphic fungus Penicillium marneffei, is an important opportunistic systemic fungal infection affecting immunocompromised individuals living in areas where penicilliosis is endemic. We have demonstrated previously that a urinary enzyme-linked immunosorbent assay (ELISA) with purified rabbit polyclonal antibody against killed whole-fission-form arthroconidia of P. marneffei was specific and highly sensitive for the diagnosis of penicilliosis. In this study, a dot blot ELISA and a latex agglutination (LA) test were developed with the same polyclonal antibody and compared with the ELISA for the detection of P. marneffei urinary antigen. Urine specimens from 37 patients with culture-proven penicilliosis and 300 controls (52 healthy subjects and 248 hospitalized patients without penicilliosis) were tested. Antigen was detected in urine from all 37 (100%) penicilliosis patients by the LA test, 35 (94.6%) penicilliosis patients by the dot blot ELISA, and 36 (97.3%) penicilliosis patients by the ELISA. False-positive results were found by the three assays for 2 (0.7%), 8 (2.7%), and 6 (2%) of 300 controls, respectively. The overall sensitivities of the diagnostic tests were as follows: dot blot ELISA, 94.6%; ELISA, 97.3%; and LA test, 100% (specificities, 97.3, 98, and 99.3%, respectively). The LA test is simple, robust, rapid, and convenient and should prove to be an important addition to the existing diagnostic tests for penicilliosis.     

Delay in diagnosis of imported Plasmodium falciparum malaria in children

The study reported here prospectively evaluated the time-to-diagnosis of imported Plasmodium falciparum malaria in children in seven French pediatric emergency departments during a 1-week period. For the 29 patients included, the mean patient, doctor and total delays were 3.1, 1.5 and 4.7 days, respectively. The late medical diagnosis for 11 patients was mainly due to the treating physician’s failure to consider malaria, despite having been informed that the child had been in an endemic area, and erroneously making a diagnosis of viral infection. The five patients who were diagnosed correctly without delay had higher mean platelet counts than the others (206,000 vs 118,541/mm³; p=0.008). The results indicate that greater awareness of the risk of malaria in returning travelers may help reduce delays in diagnosis and its consequences.     

Diagnostic efficacy of two rapid tests for detection of respiratory syncytial virus antigen.

With the availability of ribavirin therapy for serious respiratory syncytial virus (RSV) infections, rapid diagnostic tests for the detection of RSV antigen are increasingly important. Efficacies of a commercially available enzyme immunoassay (EIA) (Abbott Laboratories, North Chicago, Ill.) and a fluorescent-antibody assay (FA) were evaluated in a study involving 135 specimens from children with respiratory symptoms. A nasal wash specimen was cultured immediately on RSV-sensitive A549 cells; the nasal wash was also used for EIA. FA was performed on a nasopharyngeal swab specimen with bovine anti-RSV and anti-bovine immunoglobulin G antisera (Burroughs Wellcome Co., Research Triangle Park, N.C.). A total of 39 specimens (28%) were tissue culture positive, including 35 EIA-positive and 37 FA-positive samples (sensitivities, 90 and 95%, respectively). All 96 tissue culture-negative specimens were EIA negative (specificity, 100%); 94 of these 96 specimens were FA negative (specificity, 98%). Positive and negative predictive values for the tests were as follows: 100 and 96% for EIA, respectively, and 95 and 98% for FA, respectively. Other viruses, including influenza A virus, adenovirus, enterovirus, and herpes simplex virus, were isolated in nine cases. One adenovirus-positive specimen had a false-positive RSV FA result; all nine specimens were RSV EIA negative. Both tests performed well in our study and provide cost-effective alternatives to tissue culture. The RSV EIA, in particular, uses standard serologic techniques and equipment and does not require expertise in virology. More widespread availability of rapid diagnostic tests for RSV will hopefully result in early and appropriate use of antiviral therapy in patients at risk for serious RSV infections.     

Devices for rapid diagnosis of Malaria: evaluation of prototype assays that detect Plasmodium falciparum histidine-rich protein 2 and a Plasmodium vivax-specific antigen

The ParaSight F test was developed as a pioneer industry effort in the large-scale, process-controlled production of a device for the rapid diagnosis of malaria. This device performed well in field settings but was limited to the detection of a single malaria species, Plasmodium falciparum. The ParaSight F+V assay advanced upon the ParaSight F test format by incorporating a monoclonal antibody directed against a proprietary Plasmodium vivax-specific antigen, in addition to the antibody directed against P. falciparum histidine-rich protein 2, which was used in the ParaSight F assay. The modified assay was developed to add the capability to detect P. falciparum and P. vivax in a single-test-strip format. The present study evaluated three distinct ParaSight F+V prototypes with samples from symptomatic patients in regions of Thailand and Peru where malaria is endemic. Over a 2-year enrollment period (1998 and 1999), a total of 4,894 patients consented to participation in the study. Compared with the results for duplicate microscopic examinations of Giemsa-stained blood smears as the reference diagnostic standard, each successive prototype showed substantial improvement in performance. The final ParaSight F+V prototype, evaluated in 1999, had an overall sensitivity for detection of asexual P. falciparum parasites of 98%. The sensitivity of the device was 100% for P. falciparum densities of >500 parasites/ micro l, with a sensitivity of 83% for parasite densities of 5,000/ micro l, 92% for parasite densities of 1,001 to 5,000/ micro l, 94% for parasite densities of 501 to 1,000/ micro l, and 55% for parasite densities of 1 to 500/ micro l. The specificity for the exclusion of P. vivax was 87%. The areas under the receiver operating characteristic curves for the diagnostic performance of the assay for the detection of P. falciparum and P. vivax were 0.8907 and 0.8522, respectively. These findings indicate that assays for rapid diagnosis have the potential to enhance diagnostic capabilities in those instances in which skilled microscopy is not readily available.     

Use of rapid diagnostic tests for diagnosis of malaria in the UK

BACKGROUND: Malaria is currently diagnosed almost exclusively by microscopy in clinical laboratories. The introduction of rapid diagnostic tests (RDTs) may be useful in achieving rapid detection of malaria parasites, especially in situations where malaria is not often seen or where staff are inexperienced. AIM: To explore the use of RDT in UK laboratories. METHODS: The current use of RDTs was surveyed in UK laboratories subscribing to the United Kingdom National External Quality Assessment Scheme blood parasitology and haematology schemes. RESULTS: An overall survey response rate of 60.3% was seen. RDTs were found to be the preferred choice, either alone or in conjunction with microscopy in 31.2% of the samples examined during normal working hours and in 44.3% of the specimens examined on call. CONCLUSIONS: During on-call hours, the use of RDTs was observed to increase and RDTs changed the diagnosis in 12% of laboratories. No established protocol for RDT use was, however, observed in the UK. A protocol that needs to be validated in the laboratory setting is suggested.     

Effect of sequence variation in Plasmodium falciparum histidine- rich protein 2 on binding of specific monoclonal antibodies: Implications for rapid diagnostic tests for malaria

The ability to accurately diagnose malaria infections, particularly in settings where laboratory facilities are not well developed, is of key importance in the control of this disease. Rapid diagnostic tests (RDTs) offer great potential to address this need. Reports of significant variation in the field performance of RDTs based on the detection of Plasmodium falciparum histidine-rich protein 2 (HRP2) (PfHRP2) and of significant sequence polymorphism in PfHRP2 led us to evaluate the binding of four HRP2-specific monoclonal antibodies (MABs) to parasite proteins from geographically distinct P. falciparum isolates, define the epitopes recognized by these MABs, and relate the copy number of the epitopes to MAB reactivity. We observed a significant difference in the reactivity of the same MAB to different isolates and between different MABs tested with single isolates. When the target epitopes of three of the MABs were determined and mapped onto the peptide sequences of the field isolates, significant variability in the frequency of these epitopes was observed. These findings support the role of sequence variation as an explanation for variations in the performance of HRP2-based RDTs and point toward possible approaches to improve their diagnostic sensitivities.     

Comparison of different PCR protocols for the detection and diagnosis of Plasmodium falciparum

An assessment of differing PCR protocols for the diagnosis of Plasmodium falciparum infection was performed on samples from an area of holoendemic malaria transmission in western Burkina Faso. The PCR protocols had generally high sensitivities (>92%) and specificities (>69%), but the negative predictive values (NPV) were moderate and differed widely among the PCR protocols tested. These PCR protocols that amplified either the P. falciparum pfcrt gene or the small subunit ribosomal DNA were the most reliable diagnostic tools. However, the moderate NPV imply that more than one PCR protocol should be used for diagnosis in holoendemic areas.N. Oster and I.Z. Abdel-Aziz have contributed equally to this work.     

Comparison of two rapid streptococcal antigen detection assays with culture for diagnosis of streptococcal pharyngitis.

In this study, 801 pharyngeal specimens were cultured for group A streptococci and tested with the Biostar Strep A Optical Immunoassay (Strep A OIA). The respective sensitivities and specificities were as follows: culture, 97.1 and 100%; Strep A OIA, 91.5 and 94.8%. Of the 801 specimens, 597 were also tested with the Abbott TestPack Strep A Assay (TP-ST). For those specimens tested by all three methods, the respective sensitivities and specificities were as follows: culture, 98.1 and 100%; Strep A OIA, 92.3 and 95.4%; and TP-ST, 79.4 and 100%. The Strep A OIA is significantly more sensitive than TP-ST and compares favorably with culture.     

Evaluation & usefulness of a immunochromatographic test for rapid detection of Plasmodium falciparum infection

Rapid test (Parachek Pf) based on detection of HRP-2 protein specific to Plasmodium falciparum by immunochromatographic technique was evaluated. Prevalence of infection was 8.5%. The test was 100% sensitive & 99.5% specific on comparison with light microscopy. The test is useful for making 'on the spot' diagnosis.