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

Malaria Journal - Tập 20 Số 1 - 2021
Kim van Bergen1, Toon Stuitje2, Robert C. Akkers1, Eric Vermeer1, Rob Castel1, T. G. Mank3
1Result Laboratorium, Albert Schweitzer Hospital, Dordrecht, The Netherlands
2MRC Holland, Amsterdam, The Netherlands
3Streeklaboratorium Voor de Volksgezondheid Kennemerland, Haarlem, The Netherlands

Tóm tắt

Abstract 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.

Từ khóa


Tài liệu tham khảo

Ashley EA, Phyo AP, Woodrow CJ. Malaria. Lancet. 2018;391:608–21.

Sutherland CJ, Tanomsing N, Nolder D, Oguike M, Jennison C, Pukrittayakamee S, et al. Two nonrecombining sympatric forms of the human malaria parasite Plasmodium ovale occur globally. J Infect Dis. 2010;201:1544–50.

Anstey NM, Grigg MJ. Zoonotic malaria: the better you look, the more you find. J Infect Dis. 2019;219:679–81.

Imwong M, Madmanee W, Suwannasin K, Kunasol C, Peto TJ, Tripura R, et al. Asymptomatic natural human infections with the simian malaria parasites Plasmodium cynomolgi and Plasmodium knowlesi. J Infect Dis. 2019;219:695–702.

Lalremruata A, Magris M, Vivas-Martinez S, Koehler M, Esen M, Kempaiah P, et al. Natural infection of Plasmodium brasilianum in humans: man and monkey share quartan malaria parasites in the Venezuelan Amazon. EBioMedicine. 2015;2:1186–92.

Brasil P, Zalis MG, Pina-Costa AD, Siquira AM, Junior CB, Silva S, et al. Outbreak of human malaria caused by Plasmodium simium in the Atlantic Forest in Rio de Janeiro: a molecular epidemiological investigation. Lancet Glob Health. 2017;5:e1038–46.

Grigg MJ, Snounou G. Plasmodium simium: a Brazilian focus of anthropozoonotic vivax malaria? Lancet Glob Health. 2017;5:e961–2.

Mathison BA, Pritt BS. Update on malaria diagnostics and test utilization. J Clin Microbiol. 2017;55:2009–17.

Garrido-Cardenas JA, Gonzalez-Ceron L, Manzano-Agugliaro F, Mesa-Valle C. Plasmodium genomics: an approach for learning about and ending human malaria. Parasitol Res. 2019;118:1–27.

Kantele A, Jokiranta TS. Review of cases with the emerging fifth human malaria parasite Plasmodium knowlesi. Clin Infect Dis. 2011;52:1356–62.

Chavatte J, Tan SBH, Snounou G, Lin RTBV. Molecular characterization of misidentified Plasmodium ovale imported cases in Singapore. Malar J. 2015;14:454.

Cheaveau J, Mogollon DJ, Mohon MAN, Golassa L, Yewhalaw D, Pillai DR. Asymptomatic malaria in the clinical and public health context. Expert Rev Anti Infect Therapy. 2019;17:997–1010.

Chen I, Clarke SE, Gosling R, Hamainza B, Killeen G, Magill A, et al. "Asymptomatic” malaria: a chronic and debilitating infection that should be treated. PLoS Med. 2016;13:e1001942.

Huang F, Takala-Harrison S, Liu H, Xu J, Yang H, Adams M, et al. Prevalence of clinical and subclinical Plasmodium falciparum and Plasmodium vivax malaria in two remote rural communities on the Myanmar-China border. Am J Trop Med Hyg. 2017;97:1524–31.

Bousema T, Okell L, Felger I, Drakeley C. Asymptomatic malaria infections: detectability, transmissibility and public health relevance. Nat Rev Microbiol. 2014;12:833–40.

Cunningham JA, Thomson RM, Murphy SC, Paz Ade MDI, Ding XC, Incardona S, et al. WHO malaria nucleic acid amplification test external quality assessment scheme: results of distribution programmes one to three. Malar J. 2020;19:129.

Murphy SC, Hermsen C, Douglas AD, Edwards NJ, Petersen I, Fahle GA, et al. External quality assurance of malaria nucleic acid testing for clinical trials and eradication surveillance. PLoS ONE. 2014;9:e0097398.

Vasoo S, Pritt BS. Molecular diagnostics and parasitic disease. Clin Lab Med. 2013;33:461–503.

Padley D, Health A, Sutherland C, Chiodini P, Baylis S, Collaborative Study Group. Establishment of the 1st World Health Organization International Standard for Plasmodium falciparum DNA for nucleic acid amplification technique (NAT)-based assays. Malar J. 2008;7:139.

Chesher D. Evaluating assay precision. Clin Biochem Rev. 2008;29(Suppl 1):S23–6.

Farrugia C, Cabaret O, Botterel, Bories C, Foulet F, Costa J, et al. Cytochrome b gene quantitative PCR for diagnosing Plasmodium falciparum infection in travelers. J Clin Microbiol. 2011;49:2191–5.

Talundzic E, Ravishankar S, Nayak V, Patel DS, Olsen C, Sheth M, et al. First full draft genome sequence of Plasmodium brasilianum. Genome Announc. 2017;5:e01566-e1616.

Leblanc C, Vasse C, Minodier P, Mornand P, Naudin J, Quinet B, et al. Management and prevention of imported malaria in children. Update of the French guidelines. Med Mal Infect. 2020;50:124–214.

Lalloo D, Shingadia D, Bell D, Beeching N, Whitty C, Chiodini P, et al. UK malaria treatment guidelines 2016. J Infect. 2016;72:635–49.

WHO. Severe malaria. Trop Med Int Health. 2014;19:7–131.