Contribution of Quantitative Real-Time Polymerase Chain Reaction to Follow-Up of Visceral Leishmaniasis Patients Treated with Meglumine Antimoniate

Karim, Abdul; Chouihi, E.; Amri, Fethi; Alaya, Nissaf Ben; Raies, Ali; Mary, Charles; Bouratbine, A.

doi:10.4269/ajtmh.2009.09-0285

Cited by 16 publications

(18 citation statements)

References 9 publications

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“…In recent years, cases of therapeutic failure have been reported in the Maghreb, an area where visceral and cutaneous leishmaniasis are endemic and where meglumine antimoniate (Glucantime) is the first-line treatment (23,24).…”

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confidence: 99%

Heterogeneity of Molecular Resistance Patterns in Antimony-Resistant Field Isolates of Leishmania Species from the Western Mediterranean Area

Jeddi

Mary

Karim

et al. 2014

Antimicrob Agents Chemother

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h Antimonials remain the first-line treatment for the various manifestations of leishmaniasis in most areas where the disease is endemic, and increasing cases of therapeutic failure associated with parasite resistance have been reported. In this study, we assessed the molecular status of 47 clinical isolates of Leishmania causing visceral and cutaneous leishmaniasis from Algeria, Tunisia, and southern France. In total, we examined 14 genes that have been shown to exhibit significant variations in DNA amplification, mRNA levels, or protein expression with respect to resistance to antimonials. The gene status of each clinical isolate was assessed via qPCR and qRT-PCR. We then compared the molecular pattern against the phenotype determined via an in vitro sensitivity test of the clinical isolates against meglumine antimoniate, which is considered the reference technique. Our results demonstrate significant DNA amplification and/or RNA overexpression in 56% of the clinical isolates with the resistant phenotype. All clinical isolates that exhibited significant overexpression of at least 2 genes displayed a resistant phenotype. Among the 14 genes investigated, 10 genes displayed either significant amplification or overexpression in at least 1 clinical isolate; these genes are involved in several metabolic pathways. Moreover, various gene associations were observed depending on the clinical isolates, supporting the multifactorial nature of Leishmania resistance. Molecular resistance features were found in the 3 Leishmania species investigated (Leishmania infantum, Leishmania major, and Leishmania killicki). To our knowledge, this is the first report of the involvement of molecular resistance genes in field isolates of Leishmania major and Leishmania killicki with the resistance phenotype.

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confidence: 99%

Heterogeneity of Molecular Resistance Patterns in Antimony-Resistant Field Isolates of Leishmania Species from the Western Mediterranean Area

Jeddi

Mary

Karim

et al. 2014

Antimicrob Agents Chemother

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“…Molecular diagnosis of VL is essentially based on PCR assays. Quantitative real-time PCR (qPCR) technology, using primers designed from kinetoplast DNA (kDNA), has been successfully used on blood samples with 100% sensitivity (4,20). However, blood collection remains an invasive procedure that demands technical expertise.…”

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Diagnosis of Mediterranean Visceral Leishmaniasis by Detection of Leishmania Antibodies and Leishmania DNA in Oral Fluid Samples Collected Using an Oracol Device

et al. 2011

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Current methods for diagnosis of visceral leishmaniasis (VL) require invasive sampling procedures such as visceral aspiration and/or blood drawing. The use of diagnostic tests using oral fluid, which is easier to collect, would be more simple and practical for VL diagnosis, especially under field conditions. Oral fluids from 37 VL patients and 40 healthy controls were collected using Oracol devices. Blood samples and oral fluid specimens from both groups were analyzed by recombinant protein K39 (rK39) enzyme-linked immunosorbent assay and quantitative real-time PCR. Detection of antibodies in the oral fluid had a sensitivity of 100% and a specificity of 97.5%. Antibody levels measured in serum and oral fluid showed a significant positive correlation ( ‫؍‬ 0.655 and P ‫؍‬ 0.01). Detection of Leishmania DNA in oral fluid had a sensitivity of 94.6% and a specificity of 90%. The median parasite load estimated in blood was 133 parasites/ml (interquartile range [IR], 10 to 1,048), whereas that in oral fluid specimens was 3 parasites/ml (IR, 0.41 to 92). However, there was no significant linear relationship between parasite loads assessed in the two biological samples ( ‫؍‬ 0.31 and P ‫؍‬ 0.06). VL diagnosis based on specific antibody detection and Leishmania DNA identification using oral fluid samples was equivalent in accuracy to that using blood and therefore is promising for clinical use.

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“…They did not present with immunosuppressive diseases or risk factors of human immunodeficiency infection. VL diagnosis was suspected upon clinical signs and confirmed by both the microscopic observation of Leishmania amastigotes in Giemsa-stained bone marrow smears and real-time PCR performed on blood samples (2). Matched controls were selected among patients hospitalized in the same period for other diseases.…”

Section: Vl Patients and Controlsmentioning

confidence: 99%

“…It is generally based on the detection of Leishmania parasites in bone marrow aspirates (37,38). Recently quantitative real-time PCR technology (qPCR), using primers designed on kinetoplast DNA (kDNA), was successfully performed on blood samples with high sensitivity (2). Human VL is also diagnosed by detecting IgG that specifically binds to Leishmania antigens.…”

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confidence: 99%

Evaluation of an Enzyme-Linked Immunosorbent Assay Based on Crude Leishmania Histone Proteins for Serodiagnosis of Human Infantile Visceral Leishmaniasis

Lakhal

Mekki

Ben-Abda

et al. 2012

Clin Vaccine Immunol

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ABSTRACTHuman visceral leishmaniasis (VL) is routinely diagnosed by detecting IgG that specifically binds toLeishmaniaantigens. The enzyme-linked immunosorbent assay (ELISA) remains a widely used method. However, the biggest challenge remains the choice of antigen with the highest specificity and sensitivity. This study is aimed at assessing the diagnostic performances of crudeLeishmaniahistone (CLH) protein-based ELISAs in Mediterranean VL patients. The CLH proteins were biochemically purified from promastigote nuclear extracts. Their reactivities were analyzed by Western blotting (WB) using rabbit polyclonal antibodies againstLeishmaniarecombinant histones and sera from VL patients, respectively. Then, the diagnostic potential of CLH proteins was validated by the CLH-based ELISA using 42 infantile VL patients' sera and 70 control subjects. The CLH-based ELISA performance was compared to that of the solubleLeishmaniaantigen (SLA)- and the recombinant K39 (rK39)-based ELISAs. Analysis of the WB profile with the use of polyclonal antibodies confirmed the histone origin of low molecular mass proteins (12 to 16 kDa). All VL samples tested presented antibodies reacting against different antigen fractions; however, recognition patterns were different depending on the reactivity of each serum. CLH-based ELISA showed an excellent ability to discriminate between VL cases and healthy controls (97.6% sensitivity and 100% specificity). It had a diagnostic performance similar to that of rK39-based ELISA (97.6% sensitivity and 97.1% specificity,P= 0.5) and a better serodiagnosis accuracy than the SLA-based ELISA (85.7% sensitivity and 90% specificity,P< 0.05). Therefore, crudeLeishmaniahistone extract could be a valuable antigen for clinical use.

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Contribution of Quantitative Real-Time Polymerase Chain Reaction to Follow-Up of Visceral Leishmaniasis Patients Treated with Meglumine Antimoniate

Cited by 16 publications

References 9 publications

Heterogeneity of Molecular Resistance Patterns in Antimony-Resistant Field Isolates of Leishmania Species from the Western Mediterranean Area

Heterogeneity of Molecular Resistance Patterns in Antimony-Resistant Field Isolates of Leishmania Species from the Western Mediterranean Area

Diagnosis of Mediterranean Visceral Leishmaniasis by Detection of Leishmania Antibodies and Leishmania DNA in Oral Fluid Samples Collected Using an Oracol Device

Evaluation of an Enzyme-Linked Immunosorbent Assay Based on Crude Leishmania Histone Proteins for Serodiagnosis of Human Infantile Visceral Leishmaniasis

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