Evaluation of a novel real-time PCR assay for the detection, identification and quantification of Plasmodium species causing malaria in humans.

BACKGROUND: The entry of PCR-based techniques into malaria diagnostics has improved the sensitivity and specificity of the detection of Plasmodium infections. It has been shown that humans are regularly infected by at least six different Plasmodium species. The MC004 real-time PCR assay for malaria diagnosis is a novel single-tube assay that has been developed for the purpose of simultaneously detecting all Plasmodium species known to infect humans, and discrimination between Plasmodium falciparum, Plasmodium vivax, Plasmodium malariae, Plasmodium ovale wallikeri, Plasmodium ovale curtisi, Plasmodium knowlesi (including differentiation of three strains) and Plasmodium cynomolgi (including differentiation of three strains). Detection and identification of Plasmodium species relies on molecular beacon probe-based melting curve analysis. In addition, this assay might be used to quantify the parasitaemia of at least P. falciparum by calculating the level of parasitaemia directly from the Cq-value. METHODS: The samples used in this study comprised reference samples, patient samples, and synthetic controls. The following analytical performance characteristics of the MC004 assay were determined: analytical specificity, limit of detection, the ability to detect mixed infections, and the potential to determine the level of parasitaemia of P. falciparum, including assessment of within-run and between-run precisions. RESULTS: No false positive or false negative results were observed. The limit of detection of P. falciparum was 1 × 10-3 IU/mL (WHO standard). Mixed infections with P. falciparum and non-falciparum species were correctly identified. A calibration curve could be established to quantify the parasitaemia of at least P. falciparum. The within-run and between-run precisions were less than 20% CV at the tested parasitaemia levels of 0.09%, 0.16%, 2.15% and 27.27%. CONCLUSION: Based upon the analytical performance characteristics that were determined, the MC004 assay showed performance suitable for use in clinical settings, as well as epidemiological studies.

[Analysis of the intoxicated patient: pitfalls]. / Analyse van een patiënt met intoxicatie: valkuilen.

A 39-year-old man was admitted to the ICU after having taken 96 tablets of 500 mg acetylsalicylic acid. Although a high plasma concentration (1040 mg/l) was found, underestimation and misinterpretation of clinical signs and symptoms with decreasing salicylate plasma concentrations led to haemodialysis being postponed. One day after admission the patient suddenly died of cardiovascular collapse due to severe salicylate toxicity. Doctors should be aware that underestimating the severity of salicylate intoxication is a common and dangerous pitfall. Another patient, a 60-year-old woman, was suspected of suffering from toxic alcohol intoxication. She had marked hyperlactaemia (17 mmol/l), which could not be explained from a clinical perspective. However, ethanol was administered intravenously while awaiting the toxic alcohol test results (ethylene glycol: 1300 mg/l). The hyperlactaemia was artificial and caused by an interaction between glycolic acid and lactate on a point-of-care analyser. Doctors should consider artificial hyperlactaemia in patients with ethylene glycol intoxication. The patient recovered.