ENGLISH: Mediterranean visceral leishmaniasis is a zoonotic disease caused by Leishmania infantum and transmitted by the bites of infected female sand flies. Iran is one of the endemic areas of this disease. Dogs and canines are the major reservoir hosts of Leishmania infantum in the new and old world, including Iran. If visceral leishmaniasis is left untreated, it may result in a 90% mortality rate. The identification and elimination of infected dogs are efficient ways to control this disease. The diagnostic methods used to identify these animals cannot yield 100% detection. Therefore, in the present study, we used a multiepitope recombinant protein (PQ10) to distinguish between symptomatic and asymptomatic infections caused by Leishmania infantum in animal reservoirs (dogs). The coding sequence of PQ10 recombinant protein was sub-cloned in pET28 expression vector and was commercially synthesized by GENERAY Biotechnology, China. In the following process, sequencing was performed with plasmid-specific primers and followed by the expression, optimization of expression. The purified recombinant protein was confirmed using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blot. The efficacy of recombinant PQ10 protein was evaluated by an indirect enzyme-linked immunosorbent assay (ELISA) test using 150 dog sera (25 symptomatic positive sera, 25 asymptomatic positive sera, 80 negative sera, and 20 sera of dogs with other infectious diseases). Direct agglutination test (DAT) as the standard method was used to compare and determine specificity and sensitivity. The results indicated that the sensitivity and specificity of ELISA using infected dog sera were 94% and 86%, respectively. Positive and negative predictive values for dog sera were reported as 87. 03% and 93. 47%, respectively. This protein was able to identify 92% of asymptomatic dogs with visceral leishmaniasis. The results showed that the recombinant protein PQ10 is able to identify positive cases of canine visceral leishmaniasis, especially asymptomatic cases. FRENCH: Ré sumé : La leishmaniose viscé rale mé diterrané enne est une maladie zoonotique causé e par Leishmania infantum et transmise par les piqû res de phlé botomes femelles infecté es. L'Iran est l'une des zones où cette maladie est autochtone. Les chiens et les Canidé s sont les principaux hô tes ré servoirs de Leishmania infantum dans le nouveau et l'ancien monde, y compris l'Iran. Si la leishmaniose viscé rale n'est pas traité e, elle peut entraî ner un taux de mortalité de 90%. L'identification et l'é limination des chiens infecté s repré sentent des moyens efficaces pour contrô ler cette maladie. Les mé thodes de diagnostic utilisé es pour identifier les animaux infecté s ne peuvent pas permettre une dé tection à 100%. Par consé quent, dans cette é tude, nous avons utilisé une proté ine recombinante multiepitopique (PQ10) afin de distinguer les infections symptomatiques et asymptomatiques causé es par Leishmania infantum dans les ré servoirs animaux (chiens). La sé quence codante de la proté ine recombinante PQ10 a é té sous-cloné e dans le vecteur d'expression pET28 et a é té synthé tisé e commercialement par GENERAY Biotechnology, Chine. Dans le processus suivant, le sé quenç age a é té effectué avec des amorces spé cifiques du plasmide et suivi par l'expression et l'optimisation de l'expression. La purification de la proté ine recombinante a é té confirmé e par é lectrophorè se sur gel de sodium dodé cyl sulfate-polyacrylamide (SDS-PAGE) et Western blot. L'efficacité de la proté ine PQ10 recombinante a é té é valué e par un dosage d'immunoabsorption par enzyme (la mé thode immuno-enzymatique ELISA) utilisant 150 sé rums de chien (25 sé rums positifs symptomatiques, 25 sé rums positifs asymptomatiques, 80 sé rums né gatifs et 20 sé rums de chiens atteints d'autres maladies infectieuses). Le test d'agglutination directe (TAD) é té utilisé comme mé thode standard pour comparer et dé terminer la spé cificité et la sensibilité . Les ré sultats ont indiqué que la sensibilité et la spé cificité de l'ELISA pour identifier les sé rums de chiens infecté s é taient de 94% et 86%, respectivement. Les valeurs pré dictives positives et né gatives pour les sé rums de chiens ont é té rapporté es à 87, 03% et 93, 47%, respectivement. Cette proté ine a permis d'identifier 92% des chiens asymptomatiques atteints de leishmaniose viscé rale. Les ré sultats ont montré que l’ ELISA à base de proté ine recombinante PQ10 est capable d'identifier les cas positifs de leishmaniose viscé rale canine, en particulier les cas asymptomatiques.

Background: Iran is one of the endemic areas of Mediterranean Visceral Leishmaniasis, a disease caused by Leishmania infantum. In this work, we examined whether Proteí na quimé rica 10 (PQ10) recombinant protein is suitable for immunological diagnosis of human visceral leishmaniasis. Methods: The study was carried out in Tarbiat Modares University during 2016-2018. The coding sequence of PQ10 recombinant protein was sub-cloned in pET28 expression vector and was commercially synthesized by GENERAY Biotechnology, China. Sequencing with proper primers was done, the expression, optimization of expression and protein purification were performed, and the purified recombinant protein was confirmed by western blot. The efficacy of PQ10 for serodiagnosis was evaluated with 50 positive and 50 negative serum samples, which confirmed by the direct agglutination test and collected from individuals living in the visceral leishmaniasis endemic areas of Iran. ELISA was performed with the PQ10 recombinant protein. Results: The 95% CI sensitivity of ELISA that was evaluated with sera from naturally infected individuals was 84%. The 95% CI specificity value of the ELISA determined with sera from healthy individuals (50 serum samples) and from individuals with other infectious diseases was 82%. The 95% CI positive predictive value (PPV) and negative predictive value (NPV) were exterminated 82. 35% and 83. 67%, respectively. Conclusion: We have used a recombinant synthetic protein to improve serodiagnosis of human visceral leishmaniasis. PQ10 could be useful for diagnosis of asymptomatic cases, as well as in the early phase of infections.