Identification of Leishmania donovani in blood of experimentally infected rats by quantitative real-time PCR
Abstract
The present study has been carried out at the Department of Microbiology and Parasitology, College of Veterinary Medicine/Al-Qadisiyah University to diagnose the visceral leishmaniasis molecular technique (Quantitative real-time polymerase chain reaction). The experimental study includes fourth Wistar female rats (weighted 250 ± 2 g.) were injected by blood from an infected patient in the peritoneal cavity, after 8-10 days of experimental infection, blood samples had been collected directly from the heart in order to diagnose the infection by using quantitative real-time polymerase chain reaction. Quantitative Real-Time PCR qPCR technique was used for amplification of the conserved region in GAPDH gene that was used for detection of Leishmania donovani in blood samples of rats. It shows the Amplification of genomic DNA template concentrations of Leishmania donovani during reaction with syber dye inside the apparatus under threshold cycle, also shows the melting peak of Leishmania donovani genomic DNA template concentrations is demonstrate the specialization of Leishmania donovani genomic DNA amplification in a single peak for all samples.
References
2-Guerin PJ, Lliaro PO, Sundar S, Boelaert M, Croft SL, Desjeux P, Wasanna M, Bryceason ADM. Visceral Leishmaniasis: current status of control, diagnosis, and treatment, and a proposed research and development agenda. The Lancet infectious disease. (2002); 2: 494-501.
3-Berman J. Recent Developments in Leishmaniasis: Epidemiology, Diagnosis, and Treatment,Current infectious Disease Reports, (2005);7:33-38.
4-World Health Organization, (1990). Control of Leishmaniasis. WHO Techn. Rep, Ser.793.
5-Krobitsch S, Brandau S, Hoyer C, Schmetz C, Hübel A, Clos J. Leishmania donovani heat shock protein 100. Characterization and function in amastigote stage differentiation.J. Biol.Chem. (1998); 273:6488-6494.
6-Zhang WW, Mendez S, Ghosh A, Myler P, Ivens A, Clos J. Comparison of the A2 gene locus in Leishmania donovani and Leishmania major and its control over cutaneous infection. J. Biol .Chem. (2003); 78: 35508–35515.
7-Zhang WW, Charest H, Ghedin E, Matlashewski G. Identification and overexpression of the A2 amastigote-specific protein in Leishmania donovani. Mol. Biochem. Parasitol. (1996); 78: 79-90.
8-Charest H, Matlashewski G. Developmental Gene Expression in Leishmania donovani: Differential Cloning and Analysis of an Amastigote-Stage- Specific Gene .Mol. Cell. Biol. (1994); 14, 2975–2984.
9-Culha G, Uzun S, Ozcan K, Memisoglu HR, Chang KP. Comparison of conventional and polymerase chain reaction diagnostic techniques for leishmaniasis in the endemic region of Adana, Turkey. Int. J. Dermatol. (2006); 45:569-572.
10-Chargui N, Bastien P, Kallel K, Haouas N, Akrout FM, Masmoud A. Usefulness of PCR in the diagnosis of cutaneous leishmaniasis in Tunisia. Trans. R .Soc. Trop. Med .Hyg. (2005); 99:762-768.
11-Fraga TL, Brustoloni YM, Lima RB, Dorval MEC, Oshiro ET, Oliveira J.Polymerase chain reaction of peripheral blood as a tool for the diagnosis of visceral leishmaniasis in children. Mem. Inst. Oswaldo. Cruz. (2010); 105(3): 310-313.
12-Quaresma PF, Murta SM, Ferreira Ede C, Rocha-Lima ACD, Xavier AA, Gontijo CM. Molecular diagnosis of canine visceral leishmaniasis: identification of Leishmania species by PCR-RFLP and quantification of parasite DNA by real-time PCR. Acta. Trop. (2009); 111(3): 289-294.
13-Jassim AMH (1998). Adirect agglutination test for the diagnosis and seroepidemiological survey of visceral leishmania in basrah Governorate, Iraq. Ph. D. thesis. Collage of medicine,University of AL-Mmustansiriya. Iraq.
14-Sharpe PE, La Regina MC.The Laboratory Rat .CRC. Press. Landon, New York. (1998); PP: 115.
15-Schulz AK, Mellenthin G, Scho¨nian B, Drosten C. Detection, differentiation, and quantitation of pathogenic Leishmania organisms by a fluorescence resonance energy transfer-based real-time PCR assay. J. Clin. Microbiol. (2003); 41, 1529–1535.
16-Mary C, Faraut F, Lascombe L, Dumon H. Quantification of Leishmania infantum DNA by a real-time PCR assay with high sensitivity. J. Clin. Microbiol. (2004); 42(11): 5249-5255.
17-Wortmann GW, Romero LI, Paz HM, Ortega-Barria E, Bayard V, Hochberg LP. Real-time polymerase chain reaction diagnosis of leishmaniasis in Panama from both fresh and frozen tissue. Trans. R .Soc. Trop. Med. Hyg. (2004);98(3): 148–51.
18-Fichoux YL, Quaranta JF, Aufeuvre JP, Lelievre A, Marty P, Suffia I, Rousseau D. Occurrence of Leishmania infantum parasitemia in asymptomatic blood donors living in an area of endemicity in southern France. J. Clin. Microbiol. (1999); 37(6):1953-7.
19-Francino O, Altet L, Sanchez-Robert E, Rodriguez A, Solano-Gallego L. Advantages of real-time PCR assay for diagnosis and monitoring of canine leishmaniosis. Vet. Parasitol. (2006); 137: 214–221.
20-Rolão N, Cortes S, Rodrigues OR, Campino L. Quantification of Leishmania infantum parasites in tissue biopsies by real-time polymerase chain reaction and polymerase chain reaction-enzyme-linked immunosorbent assay. J. Parasitol. (2004); 90(5): 1150-1154.
21-Bossolasco S, Gaiera G, Olchini D, Gulletta M, Martello L, Bestetti L, Germagnoli A, Lazzarin A, Uberti-Foppa C, Cinque P. Real-time PCR assay for clinical management of human immune deficiency virus-infected patients with visceral leishmaniasis. J. Clin. Microbiol. (2003); 41: 5080-5084.
22-Zhang WW, Mendez S, Ghosh A, Myler P, Ivens A, Clos J. Comparison of the A2 gene locus in Leishmania donovani and Leishmania major and its control over cutaneous infection. J. Biol .Chem . (2003); 278: 35508-35515.
23-Mary C, Faraut F, Lascombe L, Dumon H. Quantification of Leishmania infantum DNA by a real-time PCR assay with high sensitivity. J. Clin. Microbiol. (2004); 42(11): 5249-5255.
24-Nicolas L, Prina E, Lang T, Milon G. Real-time PCR for detection and quantitation of Leishmania in mouse tissues. J. Clin. Microbiol. (2002); 40:1666–1669.
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