Contribution of specific anti-hydatid IgG subclasses in the diagnosis of echinococcosis primary infection and relapses.

BACKGROUND: Serological diagnosis of hydatid disease still faces problems of sensitivity, limiting its use to either diagnosis or post-surgical monitoring. The use of IgG subclasses seems to overcome these difficulties. The contribution of IgG subclasses was evaluated in the diagnosis of primary infested and hydatid cyst relapse patients. METHODS: A group of patients (n = 34) diagnosed for the first time with liver cystic echinococcosis (CE) and a group of patients with CE surgical recurrence were included. Enzyme-linked immunosorbent assay anti-hydatid antigens (HA) specific IgG1, 2, 3, and 4 subclasses were analyzed by ROC curves. RESULTS: ROC curve analyses demonstrated that IgG4 had the ability to discriminate between primary infested and relapsed groups whereas IgG2 was not discriminatory. The sensitivity of IgG4 was statistically higher in the relapsed cases group (97.1% versus 70.6%, p = 0.008). CONCLUSIONS: anti-HA specific IgG2 was the best marker of primary infestation whereas IgG4 was the best marker of relapse.

Diversity, Abundance and Leishmania infantum Infection Rate of Phlebotomine Sandflies in an Area with Low Incidence of Visceral Leishmaniasis in Northern Tunisia.

We report the study of sandfly Leishmania infection in an area of low incidence of visceral leishmaniasis in Tunisia. Sandflies were collected monthly using CDC light-traps set in houses and animal shelters during May-November 2016 and 2017. All males were identified at the species level. A sample of 878 females including all gravid specimens was subjected to kDNA qPCR for Leishmania detection and parasite load estimation. Leishmania species were determined by ITS1 PCR sequencing, and species identification of infected sandflies was performed by DNA barcoding. Phlebotomus perfiliewi and P. perniciosus were the dominant species during the two-year period. However, comparison of their relative abundances showed that P. perniciosus was more abundant during peaks of 2017 with longer activity duration. Real-time kDNA PCR did not detect Leishmania infection in 2016, although it identified four positive specimens (0.7%) in 2017. All four infected specimens were identified as P. perniciosus. ITS1 PCR sequencing allowed L. infantum identification in one kDNA qPCR-positive specimen. This was a P. perniciosus gravid female with a high parasite load caught during the long-lasting peak of 2017. This work highlights the usefulness of multi-seasonal studies of sandfly dynamics and kDNA qPCR in screening Leishmania infection and determining L. infantum vectors in hypo-endemic foci of human leishmaniasis.

ELISA on saliva samples for the detection of anti-hydatid cyst antibodies.

BACKGROUND: The detection of antibodies in saliva samples proved to be effective in the diagnosis of several microbial diseases. These antibodies were screened in saliva samples of patients with hydatid cysts. METHODS: Anti-hydatid fluid antigen IgG and IgA antibodies were screened in saliva and sera of patients with hydatid cysts (n=37) as well as in healthy controls (n=30) using an in-house developed immunoenzymatic assay. RESULTS: Salivary anti-hydatid fluid antigen IgG showed a sensitivity of 86.5% and a specificity of 80%. A positive correlation was observed between anti-hydatid fluid antigen IgG in saliva and in serum (r = 0.364; p = 0.02). CONCLUSIONS: The detection of anti-hydatid fluid antigen IgG antibodies in saliva using ELISA promises to be interesting for the diagnosis of cystic echinococcosis.

Temporal dynamics and Leishmania infantum infection prevalence of Phlebotomus perniciosus (Diptera, Phlebotominae) in highly endemic areas of visceral leishmaniasis in Tunisia.

Phlebotomus perniciosus is one of the major vectors of Leishmania infantum in the Mediterranean basin. The aim of this work was (i) to provide information about abundance and temporal dynamics of this Larroussius species in a hot spot area of visceral leishmaniasis in Tunisia, (ii) to detect L. infantum DNA in wild caught female sandflies and (iii) to measure Phlebotomus perniciosus infection rate throughout the active season. Sandflies were collected monthly during one year using CDC miniature light-traps in house and in animal shelters. Male specimens were identified at species level according to morphological characters. Female specimens were conserved individually for molecular study. Leishmania infection was tested by kinetoplast DNA real-time PCR and ITS-1 PCR-sequencing. Subsequent sandfly species identification of infected specimens was done by mitochondrial cytochrome b sequencing. In one year period, overall 4,441 specimens (2230 males and 2211 females) were collected. Sandfly activity started in end-April and ended in early-November. Mean sandfly density in house was significantly lower than in animal shelters (51 ± 50 versus 504 ± 460 sandflies /CDC night, p<0.05). However, a higher proportion of females was found in house (58.4% versus 49.2%, p<0.001). Based on species identification of male specimens, Phlebotomus perniciosus was the dominant species (56% of the whole male sandfly fauna, p<0.0001). It showed two peaks of density in the active season, a sharp one in early May and a higher long lasting one from end-July to end-September. DNA was extracted from 190 female specimens randomly sampled and corresponding to 96 specimens from house and 94 from animal shelters. Twenty four female sandfly were infected by Leishmania infantum. All infected specimens were recognized as Phlebotomus perniciosus. Leishmania infantum infection rate in female sandflies was 2.3 fold higher in house than in animal shelters (17.7% versus 7.4%, p<0.05). In house, estimated number of infected specimens was the highest at the end of the active season. Abundance, dynamics of density and Leishmania infantum infection prevalence of Phlebotomus perniciosus in Tunisian hot spot of visceral leishmaniasis highlight the major role of this Phlebotominae species in L. infantum transmission.

The sera from adult patients with suggestive signs of autoimmune diseases present antinuclear autoantibodies that cross-react with Leishmania infantum conserved proteins: crude Leishmania histone and Soluble Leishmania antigens [corrected].

Visceral leishmaniasis has been associated with hyper-gammaglobulinemia and antinuclear antibodies and may simulate systemic lupus erythematosus. Sera from patients with visceral leishmaniasis have been shown to strongly react against conserved proteins from the parasite, such as ribosomal and histones. Some of these proteins have also been described as immunogenic in several auto-immune syndromes, and the detection of antibodies against them is considered to be indicative of disorder in the immune system. This study aimed to assess by enzyme-linked immunosorbent assay, test routinely employed in visceral leishmaniasis diagnosis, the recognition of Crude Leishmania histone and Soluble Leishmania antigens proteins from Leishmania infantum by adult patients with suggestive signs of autoimmune diseases. Our results show that the humoral response generated during autoimmune diseases cross-reacts with the parasitic Crude Leishmania histone and Soluble Leishmania antigens. In these cases, higher precautions must be taken to confirm the presence of visceral leishmaniasis in front of positive serology in antinuclear antibodies positive sera, in order to avoid wrong diagnosis.