Comparison of Nested PCR and Conventional Analysis of Plasmodium Parasites in Kano, Nigeria
European Journal of Clinical and Biomedical Sciences
Volume 3, Issue 5, October 2017, Pages: 85-90
Received: Aug. 10, 2017; Accepted: Sep. 5, 2017; Published: Oct. 5, 2017
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Authors
Oladele Olasoji Vincent, Department of Medical Microbiology, Faculty of Medical Sciences, University of Jos, Plateau, Nigeria; Department of Educational Services, Universal Basic Education, Abuja, Nigeria
Eugene Ikeh, Department of Medical Microbiology, Faculty of Medical Sciences, University of Jos, Plateau, Nigeria
Feyi Obisakin, National Veterinary Research Institute, Jos, Plateau, Nigeria
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Abstract
Plasmodium identification represents the crucial factor in malaria diagnosis and treatment across developing countries. Conventional microscopy and the use of rapid diagnostic kits have been extensively applied towards human malaria diagnosis. Recombinant DNA techniques have been applied towards malaria diagnosis as well as in the species specific identification using Plasmodium 18s-rRNA gene. This study was undertaken amongst patients attending the Murtala Mohammed Specialist Hospital, Kano. Blood samples were collected from 350 malaria suspected patients. Microscopic analysis via Giemsa-staining revealed that 220 patients were positive for malaria. RDT analysis showed that 248 test samples were positive for Plasmodium infection. DNA products obtained from the blood samples were analyzed by nested PCR to amplify the 18S ssrRNA Plasmodium gene with genus and specific primers rPLU1/5, rPLU3/4, rVIV1/2, rFAL1/2, rMAL1/2 and rOVA1/2. Data obtained showed that 58.64% of specimens tested by microscopy were false positives while 60.62% of false positives were obtained using RDTs in comparison to nPCR which proved that on 91 out of 350 patients were infected with Plasmodium falciparum, representing 26% of tested specimen. Comparative analysis of nPCR to microscopy showed that the sensitivity and positive predictive values of the nPCR were determined as 100 and 41.36%, respectively, while against RDTs it was 100 and 39.38% respectively. nPCR was determined to be more sensitive and specific than either microscopy or RDTs. This study revealed that the accurate diagnosis of malaria by nPCR was compulsory in malaria-prone regions of Nigeria such that nPCR should be applied routinely in laboratory studies.
Keywords
Microscopy, RDTs, Nested PCR, Plasmodium falciparum, Kano
To cite this article
Oladele Olasoji Vincent, Eugene Ikeh, Feyi Obisakin, Comparison of Nested PCR and Conventional Analysis of Plasmodium Parasites in Kano, Nigeria, European Journal of Clinical and Biomedical Sciences. Vol. 3, No. 5, 2017, pp. 85-90. doi: 10.11648/j.ejcbs.20170305.11
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Copyright © 2017 Authors retain the copyright of this article.
This article is an open access article distributed under the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/) which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
References
[1]
Malik, A. M., Zaffar, N., Nadir, A., Malik, A. M., and Khan, R., (2010). Hematological Findings and Endemicity of Malaria in Gadap Region. Journal of the college of Physicians and Surgeons, 20:112-116.
[2]
WHO: World Malaria Report 2011. Geneva: World Health Organization; 2011.
[3]
WHO. World malaria report 2014. Geneva: World Health Organization; 2014.
[4]
Tukur, A. I., (2010). Temporal Variation of Malaria occurrence in Kano Municipal Local Government Area. Bayero Journal of Pure and Applied Sciences, 3(1): 132–137.
[5]
Das, L. K., and Pan, S. P., (2006) Clinical manifestation of severe form of P. falciparum malaria in Koraput district of Orissa state, India. Journal of Vector Borne Diseases, 43: 104-143.
[6]
Alonso, P. L., Brown, G., Arevalo-Herrera, M., Binka, F., Chitnis, C., Collins, F., et al., (2011). A research agenda to underpin malaria eradication. PLos Medicine, 8.
[7]
Lindblade, K. A., Steinhardt, L., Samuels, A., Kachur, S. P., and Slutsker, L., (2013). The silent threat: asymptomatic parasitemia and malaria transmission. Expert Review Anti-Infective Therapy. 11: 623–639.
[8]
Zimmerman, P. A., Mehlotra, R. K., Kasehagen, L. J., and Kazura, J. W., (2004). Why do we need to know more about mixed Plasmodium species infections in humans? Trends in Parasitology, 20: 440–447.
[9]
Muller, I., Zimmerman, P. A., and Reeder, J. C., (2007): Plasmodium malariae and Plasmodium ovale – the ‘bashful’ malaria parasites. Trends in Parasitology, 23(6): 278–283.
[10]
Mayxay, M., Pukrittayakamee, S., Newton, P. N., and White, N. J., (2004). Mixed-species malaria infections in humans. Trends in Parasitology, 20: 233–240.
[11]
Moody, A., (2002). Rapid diagnostic tests for malaria parasites. Clinical Microbiology Reviews, 15: 66–78.
[12]
Ndao, M., Bandyayera, E., Kokoskin, E., Gyorkos, T. W., MacLean, J. D., and Ward, B. J., (2004). Comparison of blood smear, antigen detection, and nested-PCR methods for screening refugees from regions where malaria is endemic after a malaria outbreak in Quebec, Canada. Journal of Clinical Microbiology, 42: 2694–2700.
[13]
Bell, D. R., Wilson, D. W., and Martin, L. B., (2005). False-positive results of a Plasmodium falciparum histidine-rich protein 2-detecting malaria rapid diagnostic test due to high sensitivity in a community with fluctuating low parasite density. American Journal of Tropical Medicine and Hygiene, 73: 199–203.
[14]
Murray, C. K., Gasser, R. A. Jr., Magill, A. J., and Miller, R. S., (2008). Update on rapid diagnostic testing for malaria. Clinical Microbiology Reviews, 21: 97–110.
[15]
Coleman, R. E., Sattabongkot, J., Promstaporm, S., Maneechai, N., Tippayachai, B., Kengluecha, A., Rachapaew, N., Zollner, G., Miller, R. S., Vaughan, J. A., Thimasarn, K., and Khuntirat, B., (2006). Comparison of PCR and microscopy for the detection of asymptomatic malaria in a Plasmodium falciparum/vivax endemic area in Thailand. Malaria Journal, 14: 121.
[16]
Okell, L. C., Ghani, A. C., Lyons, E., and Drakeley, C. J., (2009). Submicroscopic infection in Plasmodium falciparum-endemic populations: a systematic review and meta-analysis. Journal of Infectious Diseases, 200: 1509–1517.
[17]
Cheng, Q., Cunningham, J., and Gatton, M. L., (2015). Systematic Review of Sub-microscopic P. vivax infections: prevalence and determining factors. PLos Neglected Tropical Diseases, 9.
[18]
Perandin, F., Manca, N., Calderaro, A., Piccolo, G., Galati, L., Ricci, L., Medici, M. C., Arcangeletti, M. C., Snounou, G., Dettori, G., and Chezzi, C., (2004). Development of a real-time PCR assay for detection of Plasmodium falciparum, Plasmodium vivax, and Plasmodium ovale for routine clinical diagnosis. Journal of Clinical Microbiology, 42: 1214–1219.
[19]
Rougemont, M., Van Saanen, M., Sahli, R., Hinrikson, H. P., Bille, J., and Jaton, K., (2004). Detection of four Plasmodium species in blood from humans by 18S rRNA gene subunit-based and species-specific real-time PCR assays. Journal of Clinical Microbiology, 42: 5636–5643.
[20]
Taylor, B. J., Martin, K. A., Arango, E., Agudelo, O. M., Maestre, A., and Yanow, S. K., (2011). Real-time PCR detection of Plasmodium directly from whole blood and filter paper samples. Malaria Journal, 10: 244.
[21]
Silbermayr, K., Eigner, B., Duscher, G. G., Joachim, A., and Fuehrer, H. P., (2014). The detection of different Dirofilaria species using direct PCR technique. Parasitology Research Journal, 113: 513–516.
[22]
Snounou, G., and Singh, B., (2002). Nested PCR analysis of Plasmodium parasites. Methods in Molecular Medicine, 72: 189–203.
[23]
Umaru, M. L., and Uyaiabasi, G. N. (2015). Prevalence of Malaria in Patients Attending the General Hospital Makarfi, Makarfi Kaduna–State, North-Western Nigeria. American Journal of Infectious Diseases and Microbiology, 3(1): 1-5.
[24]
Paniker, C. K. J., (2007). Medical Parasitology. (6th edn.). Jaypee Brothers Medical Publishers (P) Ltd, New Delhi, India. pp 65-95.
[25]
Aduragbenro, D. A., Falade, C. O., Kotila, R. T., and Ademowo, G. O., (2007). Age as a risk factor for thrombocytopenia and anemia in children treated for acute uncomplicated falciparum malaria. Journal of Vector Borne Diseases, 44: 266-271.
[26]
Joanny, F., Lohr, S. J., Engleitner, T., Lell, B., and Mordmuller, B., (2014). Limit of blank and limit of detection of Plasmodium falciparum thick blood smear microscopy in a routine setting in Central Africa. Malaria Journal, 13: 234.
[27]
Johnston, S. P., Pieniazek, N. J., Xayavong, M. V., Slemenda, S. B., Wilkins, P. P., and da Silva, A. J., (2006). PCR as a confirmatory technique for laboratory diagnosis of malaria. Journal of Clinical Microbiology, 44: 1087–1089.
[28]
Buppan, P., Putaporntip, C., Pattanawong, U., Seethamchai, S., and Jongwutiwes, S., (2010). Comparative detection of Plasmodium vivax and Plasmodium falciparum DNA in saliva and urine samples from symptomatic malaria patients in a low endemic area. Malaria Journal, 9: 72.
[29]
Osman, M. M., Nour, B. Y., Sedig, M. F., De Bes L., Babikir, A. M., Mohamedani, A. A., and Mens, P. F., (2010). Informed decision-making before changing to RDT: a comparison of microscopy, rapid diagnostic test and molecular techniques for the diagnosis and identification of malaria parasites in Kassala, eastern Sudan. Tropical Medicine and International Health, 15: 1442–1448.
[30]
Lima, G. F., Levi, J. E., and Geraldi, M. P., (2011). Malaria diagnosis from pooled blood Samples; Comparative analysis of real-time PCR, nested PCR and Immunoassay as a platform for the molecular and serological diagnosis of malaria on a large-scale. Memorias do Instituto Oswaldo Cruz, 106(6): 691-700.
[31]
Leke, R. F., (2009). Detection of the Plasmodium falciparum antigen HRP – 2 in blood of pregnant women: implication for diagnosing placental malaria. Journal of Clinical Microbiology 37: 2992–2996.
[32]
Echeverry, D. F., Deason, N. A., Davidson, J., Makuru, V., Xiao, H., Niedbalski, J., Kern, M., Russell, T. L., Burkot, T. R., Collins, F. H., and Lobo, N. F., (2016). Malaria Journal, 15: 1-12.
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