MENA leishmaniasis.

<p>Although at least 20 <i>Leishmania spp.</i> infect humans worldwide, the primary epidemiologically relevant species in the MENA region are <i>L. major</i>, <i>L. tropica</i>, <i>L. infantum</i>, and <i>L. donovani</i>, transmitted by approximately 25 different <i>Phlebotomus spp</i>. Etiological agents of visceral leishmaniasis (VL) in MENA include <i>L. donovani</i>, <i>L. infantum</i>, and occasionally <i>L. tropica</i>. Cutaneous leishmaniasis (CL) caused by <i>L. major</i>, <i>L. tropica</i>, and <i>L. infantum</i> differ slightly in lesion presentation depending on the species. As with vector species, a variety of animal hosts have been implicated as reservoirs in the transmission of zoonotic leishmaniasis, including rodents, hyraxes, and canids. For CL caused by <i>L. major</i>, the primary cycle is zoonotic between <i>P. papatasi</i> (shown) and <i>Psammoys</i> (shown) and <i>Meriones</i> rodents. Although hyraxes have been implicated as a reservoir host for <i>L. tropica</i>, transmission is thought to be primarily anthroponotic as is the VL agent, <i>L. donovani</i>. Mediterranean VL caused by <i>L. infantum</i> is typically zoonotic where candids are the primary reservoir and man is an accidental host; however, anthroponitic cycles also have been characterized. Regardless of species or clinical manifestation, all <i>Leishmania</i> species infecting humans are transmitted by the bite of an infected sand fly. During a blood meal, metacyclic promastigotes are released by the sand fly and enter the skin of the vertebrate host. <i>Leishmania</i> parasites infect cells of the myeloid lineage, including neutrophils, followed by macrophages and dendritic cells (shown). These parasites reside within a phagolysosome where they differentiate into a dividing, aflagellated amasitogotes. Sand flies take up parasites when feeding on an infected host. Infected host cells are lysed and amastigotes differentiate into flagellated procyclic promastigotes that attach to the midgut of the sand fly vector. Subsequent development and migration towards the anterior end of the sand fly completes the cycle. Photo Credits: <i>P. papatasi</i> courtesy of Tim Gathany, Center for Disease Control Photo Services; <i>Psammomys obesus</i> from <a href="http://commons.wikimedia.org/wiki/File:Psammomys_obesus_01.jpg" target="_blank">http://commons.wikimedia.org/wiki/File:Psammomys_obesus_01.jpg</a>; rock hyrax from <a href="http://www.marietta.edu/~biol/biomes/biome_main.htm" target="_blank">http://www.marietta.edu/~biol/biomes/biome_main.htm</a>.</p

Policy recommendations for the translation of laboratory discoveries into field applications for the control of leishmanisis.

<p>Policy recommendations for the translation of laboratory discoveries into field applications for the control of leishmanisis.</p

Countries Represented.

<p>Countries Represented.</p

Leishmaniasis: Road to Translation.

<p>Leishmaniasis: Road to Translation.</p

Effect of dsRNA injection on <i>Le. major</i> infection level in <i>P. papatasi</i>.

<p>Parasite load was categorized according to the number of <i>Le. major</i> per midgut. (A) Percentage of sand flies injected with either dsCtr or dsChit1 exhibiting no infection (0 parasites), as well as light (1–1,000 parasites), moderate (1,000–10,000 parasites), or heavy (>10,000 parasites) infection at 48 h PBM. Differences are statistically significant (Chi-square, p = 0.01). (B) Percentage of sand flies injected with either dsCtr or dsChit1 exhibiting either no parasites or light infection (0–1000 parasites), or moderate infection (>1,000 parasites) at 120 h PBM. Differences are statistically significant (Fisher's exact test, p = 0.04). n: Number of flies dissected.</p

dsRNA effect on <i>PpChit1</i> RNA levels.

<p>Real-Time PCR comparing the mRNA level of <i>PpChit1</i> between flies injected with 80.5 ng (A) or 144 ng (B) of dsPpChit1 (dsChit1) or dsControl (dsCtr) double-strand RNAs. Significant <i>PpChit1</i> transcript reduction was exhibited by dsPpChit1 injected flies at 24 h (A and B), 48 h, 72 h, and 96 h PBM (A). <i>PpChit1</i> mRNA levels were normalized with the S3 housekeeping gene. Results are presented as a percent of <i>PpChit1</i> expression levels in dsPpChit1 injected flies over the mean of <i>PpChit1</i> expression levels in dsControl injected flies (considered as 100%) for each time point. The variance in PpChit1 expression in dsControl injected flies is also shown. Each dot represents <i>PpChit1</i> RNA levels in a single fly. Horizontal bars indicate mean expression level. *: Statistically significant at p<0.05.</p

Modulation of peritrophin mRNA expression upon <i>Le. major</i> infection.

<p>qRT-PCR assays depicting differences in PpPer1 (<b>A</b>), PpPer2 (<b>B</b>), and PpPer3 (<b>C</b>) mRNA levels between non-infected and <i>Le. major</i> infected midguts dissected at 24 h, 48 h, and 72 h PBM. Each dot (symbol) represents the mRNA expression levels in a single midgut whereas horizontal bars indicate mean expression levels. The cDNA encoding the S3 protein of the 40S ribosomal subunit was used as the housekeeping control gene. The mean expression of non-infected midguts was used as a standard (100%) for comparisons to the percentage of mRNA expression of <i>Le. major</i> infected midguts for each time point. NI: Non-infected. INF: <i>Le. major</i> infected. NS: Not significant. *: Statistically significant, p<0.05.</p

Persistence of bacterial infection between continuous and non-continuous feedings.

<p>Persistence of bacterial infection in sand fly larvae was determined by a qualitative assessment (presence or absence) of GFP signal using a Zeiss 510 confocal microscope. GFP positive signal observed for larvae fed for up to 48 h on bacterial lawns (continuous feeding), of for larvae fed 12 h on bacterial lawn of each bacteria, and transferred to plain LB-agar (non-continuous feeding). Larvae were collected at different time points and the midguts were dissected and prepared for confocal analyses as indicated. Results observed for the continuous and non-continuous feeding are shown as number of larvae displaying a GFP signal per total larval guts.</p><p>*As larvae in both groups fed for 12h, there is no difference between continuous and non-continuous for that time point; all larvae were grouped into the continuous feeding group.</p><p>Persistence of bacterial infection between continuous and non-continuous feedings.</p

<i>P</i>. <i>agglomerans</i> and <i>B</i>. <i>subtilis</i> infection rates.

<p><i>P</i>. <i>agglomerans</i> and <i>B</i>. <i>subtilis</i> were continuously fed to sand fly larvae. At time points indicated (left column), the infection rates in sand fly larvae were determined by a qualitative assessment (presence or absence) of GFP signal using a Zeiss 510 confocal microscope.</p><p><i>P</i>. <i>agglomerans</i> and <i>B</i>. <i>subtilis</i> infection rates.</p

dsRNA effect on <i>Le. major</i> development.

<p>Intra-thoracic injections of dsPpChit1 (80.5 ng) reduce <i>Le. major</i> load in <i>P. papatasi</i> midgut. (A). At 48 h PBM, <i>Le. major</i> density was reduced on average 46% in dsPpChit1 (dsChit1) injected compared with dsControl (dsCtr) injected. (B). <i>Le. major</i> parasites per midgut were further reduced at 120 h PBM in dsPpChit1 injected flies, reaching on average 63% reduction over the dsControl injected ones. Each dot represents parasite number in a single <i>P. papatasi</i> midgut. Horizontal bars display mean parasite numbers. n: Number of flies analyzed. *: Statistically significant at p<0.05. Graphs represent one similar result of two independent experiments.</p