Unresponsiveness to Glucantime Treatment in Iranian Cutaneous Leishmaniasis due to Drug-Resistant Leishmania tropica Parasites

BACKGROUND: Recent circumstantial evidence suggests that an increasing number of Iranian patients with cutaneous leishmaniasis are unresponsive to meglumine antimoniate (Glucantime), the first line of treatment in Iran. This study was designed to determine whether the clinical responses (healing, or non-healing) were correlated with the susceptibility of Leishmania parasites to Glucantime. METHODS AND FINDINGS: In vitro susceptibility testing was first performed on 185 isolated parasites in the intracellular mouse peritoneal macrophage model. A strong correlation between the clinical outcome and the in vitro effective concentration 50% (EC (50)) values was observed. Parasites derived from patients with non-healing lesions had EC (50) values at least 4-fold higher than parasites derived from lesions of healing patients. A selection of these strains was typed at the molecular level by pulsed-field gels and by sequencing the pteridine reductase 1 (PTR1) gene. These techniques indicated that 28 out of 31 selected strains were Leishmania tropica and that three were Leishmania major. The L. major isolates were part of a distinct pulsed-field group, and the L. tropica isolates could be classified in three related additional pulsed-field groups. For each pulsed-field karyotype, we selected sensitive and resistant parasites in which we transfected the firefly luciferase marker to assess further the in vitro susceptibility of field isolates in the monocyte cell line THP1. These determinations confirmed unequivocally that patients with non-healing lesions were infected with L. tropica parasites resistant to Glucantime. Additional characterization of the resistant isolates showed that resistance is stable and can be reversed by buthionine sulfoximine, an inhibitor of glutathione biosynthesis. CONCLUSIONS: To the authors' knowledge, this is the first report of proven resistant parasites contributing to treatment failure for cutaneous leishmaniasis and shows that primary Glucantime-resistant L. tropica field isolates are now frequent in Iran

Co-infection of Phlebotomus papatasi (Diptera: Psychodidae) gut bacteria with Leishmania major exacerbates the pathological responses of BALB/c mice

Clinical features and severity of the leishmaniasis is extremely intricate and depend on several factors, especially sand fly-derived products. Bacteria in the sand fly’s gut are a perpetual companion of Leishmania parasites. However, consequences of the concomitance of these bacteria and Leishmania parasite outside the midgut environment have not been investigated in the infection process. Herein, a needle infection model was designed to mimic transmission by sand flies, to examine differences in the onset and progression of L. major infection initiated by inoculation with “low” or “high” doses of Enterobacter cloacae and Bacillus subtilis bacteria. The results showed an alteration in the local expression of pro- and anti-inflammatory cytokines in mice receiving different inoculations of bacteria. Simultaneous injection of two bacteria with Leishmania parasites in the low-dose group caused greater thickness of ear pinna and enhanced tissue chronic inflammatory cells, as well as resulted in multifold increase in the expression of IL-4 and IL-1β and a decrease in the iNOS expression, without changing the L. major burden. Despite advances in scientific breakthroughs, scant survey has investigated the interaction between micro and macro levels of organization of leishmaniasis that ranges from the cellular to macro ecosystem levels, giving rise to the spread and persistence of the disease in a region. Our findings provide new insight into using the potential of the vector-derived microbiota in modulating the vertebrate immune system for the benefit of the host or recommend the use of appropriate antibiotics along with antileishmanial medicines

Molecular Examination of Trichomonas vaginalis Infection and Risk of Prostate Cancer in the Biopsy of Patients with Different Prostate Lesions

BACKGROUND፡ Trichomoniasis is a sexually transmitted infectious disease caused by a flagellated protozoa, Trichomonas vaginalis (T.vaginalis) and is often asymptomatic in men. Benign prostatic hyperplasia (BPH) and prostate cancer (PCA) are the most common urological diseases in the elderly. Scientists have proposed various factors which trigger prostate cancer, including sexually transmitted diseases. Thus, this study aimed to evaluate the potential role of T. vaginalis as a risk factor for various prostate lesions such as hyperplasia and prostate cancer.METHODS: A total of 250 paraffin-embedded of different prostate lesion biopsies were analyzed by Polymerase Chain Reaction (PCR) using the beta-tubulin gene for identifying T. vaginalis.RESULT: All 250 pathologic specimens were negative for this parasite by using PCR technique.CONCLUSION: It seems that T. vaginalis may have not had a causative role for different prostate lesions and it seems proposed PCR technique is an insufficient method to find the parasite in paraffin-embedded tissues. Therefore, other diagnostic techniques to identify the parasite in biopsy samples are suggested

Phylogenetic Analysis of <i>Leishmania</i> Isolates Responsible for ACL by Comparing the Sequences of <i>PTR1</i>

<p>The <i>PTR1</i> gene was sequenced in 31 Iranian isolates and control L. major cells (LV39; Friedlin), L. donovani 1S2D, L. infantum (MHOM/67/ITMAP-263), L. tropica ATCC50129, and L. tarentolae TarII. Strain 848 is identical, as determined by RAPD-PCR, to L. tropica MHOM/IR/99, and strain 749 is identical to L. major MHOM/IR/75/ER. Strains 444 and 454 were <i>Leishmania</i> parasites isolated from dogs in Iran. The sequences were compared and clustered using the Molecular Evolutionary Genetics Analysis (MEGA3) software. </p

Karyotypes of Iranian <i>Leishmania</i> Isolates as Determined by PFGE

<p> <i>Leishmania</i> cells lysed in situ in agarose blocks were run in agarose gels, and chromosome-sized bands were resolved by CHEF electrophoresis. I, II, III, and IV represent the various karyotypes, and cardinal numbers represent the identification number of <i>Leishmania</i> isolates (see <a href="http://www.plosmedicine.org/article/info:doi/10.1371/journal.pmed.0030162#pmed-0030162-t002" target="_blank">Table 2</a>). M indicates S. cerevisiae molecular-weight maker (BioRad). </p

Reversal of Antimony Resistance in Intracellular ACL Strains Using BSO

<p>The glutathione-specific inhibitor BSO (5 mM) abrogated resistance in the Glucantime-resistant strains 439, 670, 827, and 878. Only the result for strain 827 (expressing the firefly luciferase) is shown, which represents an average of two duplicate experiments. Open squares indicate strain 827; filled squares indicate strain 827 and BSO.</p

Treatment Outcome and Parasite Isolation

<p>Parasites were isolated from 185 patients with suspected ACL prior to treatment. Patients were included in three forks of treatment, and 20 patients did not respond to therapy. Parasites from these nonresponders were also isolated.</p

Parasite Susceptibility to Glucantime in the THP1 Cell Line

<p>Parasites expressing the firefly luciferase were used to infect THP1, and the levels of parasites surviving Glucantime treatment were calculated by measuring luciferase activity expressed as percentage relative light unit (RLU %). The average of two independent experiments performed in duplicate is shown. Open circles indicate strain 175 (sensitive); open triangles indicate strain 467 (intermediate resistance); and open squares indicate strain 827 (high resistance).</p