Table 2. Data about positive sheep heart samples.

<p>ND: not done.</p

Electrophoretic pattern of the direct PCR on sheep hearts.

<p>C- corresponds to negative control of PCR; MW: 100bp ladder; 1–9: amplification of different sheep isolates and C+ corresponds to the positive control of the reaction.</p

Table 1. Summary of antibodies prevalence.

<p>Ni: Initial number of tested serums; N+: Number of sera with positive reaction; N–: Number of sera with negative reaction; %: percentage of seroprevalence.</p

SplitsTree network of toxoplasmic strains at the marker SAG3.

<p>RH, PRU and CEP represent reference strains of type I, II and III respectively. TgUgCh: strains isolated form free-ranging chicken in Uganda. LA, PL, LCR: Tunisian clinical strains. ShrTgTn: strains isolated from sheep in Tunisia. CSe and CSd: atypical strains.</p

Summary of strains polymorphism at SAG3 marker.

<p>Summary of strains polymorphism at SAG3 marker.</p

Neighbor-joining tree based on 279 aligned base pairs of the cytochrome b mitochondrial DNA (mtDNA).

<p>The sequences from sandflies collected during this study (KHF8 and KHF12) were compared to Mediterranean haplotypes of <i>P</i>. <i>perniciosus</i> and <i>P</i>. <i>longicuspis</i> available in GenBank. The percent bootstrap values are indicated on the branches.</p

<i>Leishmania</i> identification by kDNA real time PCR.

<p>A: Standard curve obtained from serial dilutions of <i>Leishmania</i> DNA expressed as the number of parasites per reaction tube. The standard curve was established from <i>Leishmania</i> DNA extracted from 10⁶ <i>L</i>. <i>infantum</i> promastigotes. One μl of serial dilutions, ranging from 1000 (P1) to 0.01 parasites (P6) was introduced into reaction tubes. P5 (0.01 parasites per μl) showed a Ct = 32.8. B: Real time PCR amplification curves showing qPCR positive sandfly female specimen (F) as well as negative sandfly male specimen (M). Amplification plots obtained from the serial dilution of <i>Leishmania</i> DNA (P1 to P6) are also shown.</p

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

<div><p><i>Phlebotomus perniciosus</i> is one of the major vectors of <i>Leishmania infantum</i> in the Mediterranean basin. The aim of this work was (i) to provide information about abundance and temporal dynamics of this <i>Larroussius</i> species in a hot spot area of visceral leishmaniasis in Tunisia, (ii) to detect <i>L</i>. <i>infantum</i> DNA in wild caught female sandflies and (iii) to measure <i>Phlebotomus perniciosus</i> 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. <i>Leishmania</i> 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 <i>versus</i> 504 ± 460 sandflies /CDC night, p<0.05). However, a higher proportion of females was found in house (58.4% <i>versus</i> 49.2%, p<0.001). Based on species identification of male specimens, <i>Phlebotomus perniciosus</i> 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 <i>Leishmania infantum</i>. All infected specimens were recognized as <i>Phlebotomus perniciosus</i>. <i>Leishmania infantum</i> 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 <i>Leishmania infantum</i> infection prevalence of <i>Phlebotomus perniciosus</i> in Tunisian hot spot of visceral leishmaniasis highlight the major role of this <i>Phlebotominae</i> species in <i>L</i>. <i>infantum</i> transmission.</p></div

Densities of total sandfly and <i>Phlebotomus perniciosus</i> male specimens according to period of capture.

<p>Densities of total sandfly and <i>Phlebotomus perniciosus</i> male specimens according to period of capture.</p

<i>Leishmania</i> infection in phlebotomine female specimens.

<p><i>Leishmania</i> infection in phlebotomine female specimens.</p