Zoonotic Cutaneous Leishmaniasis Research Articles

Susceptibility of Phlebotomus papatasi (Diptera: Psychodidae) against DDT and Deltamethrin in an Endemic Focus of Zoonotic Cutaneous Leishmaniasis in Iran

Background: Phlebotomus papatasi (Diptera: Psychodidae) is the main vector of zoonotic cutaneous leishmaniasis (ZCL) in Iran. The nonstandard use of pesticides against pests, particularly in agriculture, indirectly has caused the de­velopment of resistance and, consequently, the threat of control measures in ZCL endemic areas. Up to 2023, several reports of resistance in Ph. papatasi have been declared in the Old World. The purpose of this study was to measure the lethal time (LT50 and LT90) of Ph. papatasi sand flies in the ZCL endemic center of Esfahan to DDT and deltamethrin insecticides.
 Methods: Sand flies were collected in Borkhar and were tested using WHO adult mosquito test kit against DDT 4% and deltamethrin 0.0002%. The sand fly’s survival was recorded during exposure time in 225, 450|, 900, 1800, and 3600-seconds’ intervals for DDT and Deltamethrin and they were allowed to recover for 24 hours. Then LT50 and LT90 were analyzed using probit software. Phlebotomus papatasi were identified using morphological keys and other sand flies’ species were excluded from the analysis.
 Results: The insecticide against female Ph. papatasi revealed hundred percent mortality when exposed to DDT 4% and deltamethrin 0.0002%. The LT50 and LT90 were 19.32 and 22.74 minutes for DDT 4% and 39.92 and 51.33 minutes for deltamethrin 0.0002% respectively.
 Conclusion: Results of this study revealed that Ph. papatasi is still susceptible to DDT and deltamethrin. This data pro­vides valuable knowledge to implement effective control strategies against ZCL main vector and help to manage insec­ticide resistance in the region.

Transcriptional signatures in human macrophage-like cells infected by Leishmania infantum, Leishmania major and Leishmania tropica.

In the Mediterranean basin, three Leishmania species have been identified: L. infantum, L. major and L. tropica, causing zoonotic visceral leishmaniasis (VL), zoonotic cutaneous leishmaniasis (CL) and anthroponotic CL, respectively. Despite animal models and genomic/transcriptomic studies provided important insights, the pathogenic determinants modulating the development of VL and CL are still poorly understood. This work aimed to identify host transcriptional signatures shared by cells infected with L. infantum, L. major, and L. tropica, as well as specific transcriptional signatures elicited by parasites causing VL (i.e., L. infantum) and parasites involved in CL (i.e., L. major, L. tropica). U937 cells differentiated into macrophage-like cells were infected with L. infantum, L. major and L. tropica for 24h and 48h, and total RNA was extracted. RNA sequencing, performed on an Illumina NovaSeq 6000 platform, was used to evaluate the transcriptional signatures of infected cells with respect to non-infected cells at both time points. The EdgeR package was used to identify differentially expressed genes (fold change > 2 and FDR-adjusted p-values < 0.05). Then, functional enrichment analysis was employed to identify the enriched ontology terms in which these genes are involved. At 24h post-infection, a common signature of 463 dysregulated genes shared among all infection conditions was recognized, while at 48h post-infection the common signature was reduced to 120 genes. Aside from a common transcriptional response, we evidenced different upregulated functional pathways characterizing L. infantum-infected cells, such as VEGFA-VEGFR2 and NFE2L2-related pathways, indicating vascular remodeling and reduction of oxidative stress as potentially important factors for visceralization. The identification of pathways elicited by parasites causing VL or CL could lead to new therapeutic strategies for leishmaniasis, combining the canonical anti-leishmania compounds with host-directed therapy.

Application of Rodenticides for the Control of Zoonotic Cutaneous Leishmaniasis in Iran: A Systematic Review of the Literature

Application of Rodenticides for the Control of Zoonotic Cutaneous Leishmaniasis in Iran: A Systematic Review of the Literature

Leishmania major Infection in Synanthropic Rodents: Evidence for the Urbanization of Zoonotic Cutaneous Leishmaniasis (ZCL) in Southern Iran.

Cutaneous leishmaniasis is of particular importance in southern Iran. This study aimed to investigate the infection of rodents with Leishmania major in an urban area of Fars Province, located in southern Iran. Rodents were trapped and samples from the liver, spleen, and skin were collected. Impression smears were prepared from these tissues and any skin lesions and were examined microscopically. In addition, a portion of the samples were preserved for subsequent DNA extraction. A total of 41 rodents belonging to three species were caught from 10 trapping stations in gardens or houses within the area. The caught rodent species were Rattus rattus (n = 25, 60.97%), Mus musculus (n = 15, 36.58%), and Meriones persicus (n = 1, 2.5%). Leishmania amastigotes were seen in the spleen tissue smear of 6 (2.43%) of the rodents, including 4 of R. rattus and 2 of M. musculus. Skin lesions were observed on the muzzles of two R. rattus and one M. musculus. Samples taken from these lesions tested positive for Leishmania infection. Leishmania DNA was detected in 18 (43.9%) rodents, including 11 R. rattus, 6 M. musculus, and one M. persicus, based on DNA sequencing of the ITS2 gene and PCR of the kDNA. Phylogenetic reconstruction revealed that the parasite infecting the rodents was L. major. The detection of Leishmania infection in these rodents in urban areas raises concerns about the urbanization of cutaneous leishmaniasis caused by L. major. This urbanization poses unique challenges for control and prevention efforts.

Evaluation of the Entomopathogenic Fungus Beauveria bassiana on Different Stages of Phlebotomus papatasi (Diptera: Psychodidae), Vector of Zoonotic Cutaneous Leishmaniasis in Iran

Background: Zoonotic cutaneous leishmaniasis is a major public health problem in Iran with the main vector of Phlebotomus papatasi. The use of entomopathogenic fungi for biological control of the vector is a potential substitute for the current methods which are being used. The purpose of the current study was to assess the virulence of two local isolates of Beauveria bassiana (OZ2 and TV) against Ph. papatasi.&#x0D; Methods: To perform the bioassay test, fungal suspensions were applied for every stage of the sand fly life cycle. The mortality rate, longevity, and number of eggs laid were determined. Also, the probability of fungal survival on the sur­face of rodent’s body was assessed.&#x0D; Results: The longevity of infected adult sand flies with both isolates of B. bassiana was significantly lower (P&lt; 0.05) in comparison to the negative control. The estimated Lethal concentration 50 (LC50) values for adult female and male sand flies treated with OZ2 isolate were 1.4×106 and 2.2×107 conidia/ml, respectively, while they were 6.8×106 and 2.3×108 conidia/ml for TV iso­late, respectively. Both isolates of B. bassiana exhibited nonsignificant mortality rates in sand fly larvae and pupae and fecundity rate (P&gt; 0.05). According to our findings for both isolates, the fungus continued to spread throughout the sur­face of the rodent's body for 144 hours after spraying.&#x0D; Conclusion: The current study demonstrated that both isolates of B. bassiana have considerable biological control ca­pacity against adult sand flies.

Evaluation of the in vitro Antileishmanial Activity of Chitosan-Coated Nanoemulsions of Ozonated Olive Oil

ABSTRACT Leishmaniasis is a neglected parasitic disease. Numerous studies suggested that nanoemulsions could improve drug delivery in the treatment of zoonotic cutaneous leishmaniasis. This study aims to determine the effect of chitosan-coated nanoemulsions of ozonated olive oil on Leishmania major promastigote/amastigote in vitro. Nanoemulsions (NEs) were prepared using a sonicator bath at 37 °C for one hour at 1000 W, followed by 24 hours of stirring at pH 5.5. The NE was then supplemented with chitosan and stirred all night long at a low speed. Droplet size, zeta potential, in vitro drug release, and cytotoxicity were examined to optimize the chitosan nanoemulsions (CNE) and chitosan ozonated nanoemulsions (CONE). CONE and CNE were found to have droplet sizes and zeta potentials of 16.86 nm, 19.03 nm, 19.7 mV, and 19.3 mV, respectively. Additionally, the final CONE’s drug loading and encapsulation efficiency were determined to be 100% and 3.21%, respectively. The lower doses of CONE had no cytotoxic effects on macrophages. Additionally, it has the ability to inhibit L. major promastigote/amastigote growth. Therefore, the lower doses of the CONE formulation can inhibit the growth of promastigote and amastigote of L. major in vitro while not being toxic to macrophages.

Geographical and Molecular Analysis of Haplotype Variations in Leishmania major Among Infected Iranian Phlebotomus papatasi.

Leishmania major is main causative agent and Phlebotomus papatasi is only proven vector of Zoonotic Cutaneous Leishmaniasis (ZCL) in Iran. Human leishmaniasis is mostly susceptible to climatic conditions and molecular variations of Leishmania parasites within sandflies. L. major was analyzed based on geographical, environmental, climatic changes and haplotype variations within P. papatasi. Molecular tools and different geographical aspects were employed using Arc-GIS software for mapping the geographic distribution of samples and other statistics tests. Fragments of ITS-rDNA, k-DNA, and microsatellite genes of Leishmania were used for PCR, RFLP, sequencing, and phylogenetic analyses. Totally 81 out of 1083 female P. papatasi were detected with Leishmania parasites: 70 and five were L. major and L. turanica, respectively. Golestan and Fars provinces had the highest (13.64%) and lowest (4.55%) infection rates, respectively. The infection rate among female P. papatasi collected from gerbil burrows was significantly higher (15.15%) than animal shelters, yards, and inside houses (4.48%) (P < 0.0%). Microsatellite was more sensitive (22.72%) than k-DNA (18.8%) and ITS-rDNA (7.48%). More molecular variations of L. major were found in Isfahan province. Arc-GIS software and other statistics tests were employed to find Leishmania positive and haplotype variations among sand flies. Geographical situations, altitude, climate, precipitation, humidity, temperature, urbanization, migrations, regional divergences, deforestation, global warming, genome instability, ecology, and biology of the sand flies intrinsically, and the reservoir hosts and neighboring infected locations could be reasons for increasing or decreasing the rate ofLeishmaniainfection and haplotype variations.

A prospective longitudinal study on the elimination trend of rural cutaneous leishmaniasis in southeastern Iran: Climate change, population displacement, and agricultural transition from 1991 to 2021

Leishmaniasis is a complex disease. Any change in weather conditions affects the humans' social and agricultural expansion and, consequently, the parasite's life cycle in terms of ecology, biodiversity, social stigma, and exclusion. This population-based prospective longitudinal investigation was conducted between 1991 and 2021 in a well-defined CL (cutaneous leishmaniasis) focus in Bam County, southeastern Iran. A robust health clinic and health surveillance system were responsible for the ongoing systematic documentation, detection, identification, and management of CL cases. The exponential smoothing method via the state space model was used in the univariate time series. The TTR, smooth, and forecast packages were used in R software. Landsat satellite images from 1991, 2001, 2011, and 2021 were employed in the physical development. During this period, the temperature increased while the rainfall and humidity decreased. The findings showed a downward trend in the standardized drought index. Also, the results showed that climate warming and ecological changes profoundly affected the area's agricultural patterns and topographical features. Furthermore, the last three decades witnessed an elimination trend for zoonotic CL (ZCL) and the predominance of anthroponotic CL (ACL). The present findings showed that the critical factors in the predominance of ACL and elimination of ZCL were rising temperature, drought, migration, unplanned urbanization, earthquake, and agrarian reform. The wall-enclosed palm tree gardens excluded the primary ZCL reservoir host. They controlled the disease while providing suitable conditions for the emergence/re-emergence of ACL in the newly established settlements and the unplanned ecozone. Therefore, robust health infrastructures, sustained financial support, and evidence-based research studies are crucial to facilitating the necessary surveillance, monitoring, and evaluation to control and eliminate the disease.

Diversity and dissemination of viruses in pathogenic protozoa

Viruses are the most abundant biological entities on Earth and play a significant role in the evolution of many organisms and ecosystems. In pathogenic protozoa, the presence of viruses has been linked to an increased risk of treatment failure and severe clinical outcome. Here, we studied the molecular epidemiology of the zoonotic disease cutaneous leishmaniasis in Peru and Bolivia through a joint evolutionary analysis of Leishmania braziliensis and their dsRNA Leishmania virus 1. We show that parasite populations circulate in tropical rainforests and are associated with single viral lineages that appear in low prevalence. In contrast, groups of hybrid parasites are geographically and ecologically more dispersed and associated with an increased prevalence, diversity and spread of viruses. Our results suggest that parasite gene flow and hybridization increased the frequency of parasite-virus symbioses, a process that may change the epidemiology of leishmaniasis in the region.

A Prospective cohort study of zoonotic cutaneous leishmaniasis in tunisia: Clinical and Immunological features and immune correlates of protection.

This study aimed to define immunological markers of exposure to L. major parasites and identify correlates of protection against infection. We analyzed a cohort of 790 individuals at risk of developing ZCL living in endemic areas with varying L. major infection prevalence. One area had a high infection prevalence indicated by high proportions of leishmanin skin test (LST) positive subjects, while the other areas were recent foci with lower infection prevalence. Blood samples were collected before the transmission season to measure Interferon gamma (IFN-γ), Interleukin 10 (IL-10), and Granzyme B (GrB) levels in response to parasite stimulation in peripheral blood mononuclear cells. A one-year follow-up period involved active detection of new ZCL cases to estimate disease incidence after a transmission season and identify immune correlates of protection. The study population showed heterogeneity in parasite contact, evident from specific scars and/or positive LST results, significantly higher in the old focus compared to recent foci. IFN-γ and GrB were markers of parasite exposure and reliable indicators of immunity to L. major. Positive correlations were observed between IFN-γ/IL-10 and GrB/IL-10 ratios and LST results. Unexpectedly, only 29 new ZCL cases (4%) appeared after a transmission season, with 27 cases reported in recent foci and 2 in the oldest focus. Our findings indicate that individuals in L. major endemic areas are likely to develop ZCL regardless of their LST status. We showed that high pre-transmission season levels of IFN-γ and GrB produced by PBMC, along with a high IFN-γ/IL-10 ratio, were associated with protection. This study on a large cohort at risk of ZCL confirmed IFN-γ and GrB as protective factors against the disease. A high IFN-γ/IL-10 ratio, but not GrB/IL-10 ratio was associated with resistance. These results are valuable for developing and evaluating of a vaccine against human leishmaniasis.

Field Efficacy of Topical Nano-Liposomal Amphotericin B (Sina Ampholeish®) Alone or in Combination with Glucantime® and Cryotherapy on Human Cutaneous Leishmaniasis.

Cutaneous leishmaniasis (CL) is a parasitic disease that presents a broad spectrum of clinical features. Treatment of CL is problematic. We aimed to compare the field therapeutic efficacy of topical nanoliposomes containing 0.4% amphotericin B (Nano Lip-AmB) alone and in combination with cryotherapy and/or Glucantime® on human CL in the endemic areas of Iran. This retrospective study was performed based on the results of using Nano Lip-AmB alone or with Glucantime® and/or cryotherapy in the treatment of zoonotic cutaneous leishmaniasis (ZCL) in patients referred to health centers of Isfahan, Golestan and Ilam Provinces of Iran as endemic foci of ZCL caused by Leishmania major besides Mashhad and Bam cities as endemic foci of anthroponotic cutaneous leishmaniasis (ACL) caused by with L. tropica. Two hundred and seventy-eight patients with CL were included in the current study. All of the patients (100%) who received Nano Lip-AmB alone or in combination with Glucantime® and/or cryotherapy based on guideline of Iranian national committee for the treatment of CL. Two patients with 7 skin lesions, who was resident in ACL endemic area and received Nano Lip-AmB plus Glucantime® and another patient was a resident of ZCL endemic area and received Nano Lip-AmB plus cryotherapy showed clinical relapses after treatment. Sina Ampholeish® in combination with other standard protocols of treatment of CL is well tolerated and with acceptable clinical efficacy rate.

Association Between Wolbachia Infection and Susceptibility to Deltamethrin Insecticide in Phlebotomus papatasi (Diptera: Psychodidae), the Main Vector of Zoonotic Cutaneous Leishmaniasis.

Background: Phlebotomus papatasi (Diptera: Psychodidae) is the main vector of zoonotic cutaneous leishmaniasis. Wolbachia is a symbiotic alphaproteobacteria of arthropods that can be involved in susceptibility or resistance. This study aimed to investigate the relationship between Wolbachia and Deltamethrin susceptibility/resistance in Ph. papatasi. Deltamethrin filter papers (0.00002%) were used to test sand fly field collected from southern Iran. After the test, PCR amplification of the Wolbachia surface protein gene (wsp) was used to measure Wolbachia infection rate in the killed, surviving, and control groups. Result: The rates of infection by Wolbachia strain (wPap, super group A) differed between killed (susceptible) and surviving (resistant) Ph. papatasi specimens. The rate of Wolbachia infection in susceptible individuals was more than twice (2.3) (39% vs. 17%) in resistant individuals with the same genetic background. This difference was highly significant (p < 0.001), indicating a positive association between Wolbachia infection and susceptibility to Deltamethrin. In addition, the results showed that Deltamethrin can act as a PCR inhibitor during detection of Wolbachia in Ph. papatasi. Conclusion: Results of this study show that Wolbachia is associated with Deltamethrin susceptibility level in Ph. papatasi. Also, as Deltamethrin has been identified as a PCR inhibitor, great care must be taken in interpreting Wolbachia infection status in infected populations. The results of this study may provide information for a better understanding of the host-symbiont relationship, as well as application of host symbiosis in pest management.

Antiparasitic effect of Farnesol against Leishmania major: A rationale from in vitro and in silico investigations.

Leishmaniasis is a vector-borne parasitic infection caused by the infective bite of female Phlebotomine sandflies. Treatment of leishmaniasis by conventional synthetic compounds is met by challenges pertaining to adverse effects which call for the discovery of newer anti-leishmanial molecules. This study was performed to evaluate the effect and modes of action of a sesquiterpene alcoholic molecule Farnesol on Leishmania major, the causative agent of Zoonotic CL. The cytotoxic effect of Farnesol against L.major promastigotes, amastigotes and macrophages was assessed by MTT test and counting. The IC50 on promastigotes by Farnesol on L.major was also evaluated by flow cytometry. In the findings, promastigotes were reduced at 167μM. The mean numbers of L.major amastigotes in macrophages were significantly decreased on exposure to Farnesol at 172μM. In addition, Farnesol induced significant apoptosis dose-dependent on L.major promastigotes. In silico protein-ligand_binding analyses indicated the effect of Farnesol in perturbation of the ergosterol synthesis pathway of Leishmania with attributes suggesting inhibition of Lanosterol-α-demethylase, the terminal enzyme of ergosterol synthesis machinery. Findings from flow cytometry reveal the role of Farnesol in apoptosis-induced killing in promastigotes. Farnesol was effective at very lower concentrations when compared to Paromomycin. Further studies are crucial to evaluate the therapeutic potential of Farnesol alone or in combination with other conventional drugs in animal models.

Molecular Identification and Phylogenetic Analysis of Leishmania major Isolated from Zoonotic Cutaneous Leishmaniasis Foci in Northeast Iran.

Epidemiological studies, classification and genetic studies of Leishmania species are effective in treatment, control and prevention in endemic areas. We aimed to investigate the genetic diversity and phylogeny of Leishmania in Zoonotic foci located in northeastern Iran using nagt gene for the first time. DNA of 100 confirmed positive slides collected from the health centers of Sarkhes, Darghez, Fariman, Esfarayen, and Sabzevar were extracted during 2020-2021. The partial sequence of kDNA was amplified to identify the species. Twenty-five DNA samples were randomly subjected to amplify by nagt gene primes and were sequenced. The sequences were aligned with reference sequences in National Center for Biotechnology Information (NCBI). Then, the genetic similarities of the sequences were checked using Clustalx2.1 software and the phylogenetic tree was drawn by Mega 7 software. All the positive samples were diagnosed as L. major. Approximately, half of the sequences of species were similar to two reference genes JX103550.1:404-712 L. major Esfahan and KX759012.1:568-807 L. Major Ilam (more than 90% similarity). According to the results of the phylogeny tree, the closest genotype to our study samples was JX103550.1:404-712 L. major Esfahan. The most causative agent CL in these areas was L. major. The genetic diversity of L. major was high such as other zoonotic foci in Iran. Due to the high similarity of the strains in the study areas with the strains of Isfahan and Ilam, similar control and prevention methods is suggested in these areas.

Canine Leishmania spp. infection in two distinct foci of visceral and cutaneous leishmaniasis in Tunisia.

Visceral leishmaniasis (VL) and zoonotic cutaneous leishmaniasis (ZCL) caused by Leishmania (L.) infantum and L. major, respectively, are endemic in Tunisia. The aim of the study was to assess canine Leishmania spp. infection prevalence as well as to identify the Leishmania species involved in two well-documented and geographically distinct VL and ZCL foci. One hundred seventy-six dogs were randomly recruited in the VL focus of Sbikha-Zaghouan (n = 100) and the ZCL focus of Echrarda-Nasrallah (n = 76). Physical examination and blood collection were systemically performed. Needle aspiration was done in case of lymph node (LN) enlargement. All sera were tested by ELISA. kDNA RT-PCR was performed on DNA extracts from (i) buffy coats of seropositive dogs and (ii) LN aspirates. Leishmania species identification was done by ITS1 PCR-sequencing. Thirty-three dogs (18.8%) were infected by Leishmania; 30 having anti-Leishmania antibodies and 3 were seronegative dogs with Leishmania DNA in LN aspirates. Prevalence of infection was significantly higher in VL foci than in ZCL foci (27% versus 7.9%, p = 0.002). Leishmania species was identified in 11 dogs and corresponded to L. infantum. Combination of serology and qPCR on LN aspirates seems to be the best option for canine leishmaniasis diagnosis. Infection is more frequent in VL foci and L. infantum is the only identified species.

Cutaneous Leishmaniasis: A clinicoepidemiological Study in Al Muthanna Governorate/Iraq

Background: Leishmaniasis is an endemic disease in Iraq. There are three types of leishmaniasis; cutaneous, mucocutaneous, and visceral form. Cutaneous leishmaniasis (CL) is the most common type that causes skin sores. In Iraq, CL has 2 presentations, zoonotic cutaneous leishmaniasis which is caused by L. major, and anthroponotic cutaneous leishmaniasis caused by L. Tropica. Leishmaniasis, is transmitted by the bite of infected female phlebotomine sandflies. Objective: We aimed to verify the clinical and epidemiology of cutaneous leishmaniasis in the Al Muthanna governorate. Materials and methods: A prospective cross-sectional descriptive study was carried out in the outpatient dermatological clinic in Al Hussein Teaching Hospital during the period from the 1st of December 2021 to the 1st of April 2022 in Al Muthanna governorate, Iraq. Patients with CL of any age and both sexes were enrolled in the current investigation. Detailed information regarding the age, gender, residence, duration, site, number, and type (wet or dry) of the lesion was recorded for every participant. Results: Two hundred fifty patients were diagnosed clinically as having CL. Males represented 132 (52.8%) patients. The majority of the cases were found in the age group 0-10 (n = 174, 69.6%) patients. Eighty percent (n = 200) of the participants were from rural areas. Most of the cases were presented in less than two months (n = 164, 65.6 %). Most of the lesions affected the face (n = 89, 35.6%), and the least was seen in the upper and lower limbs (n = 9, 3.6 %). The majority of the cases had a single lesion (n = 164, 65.6%) and of the dry type (n = 178, 71.2%). Conclusion: CL is an endemic dermatological disease in the Al Muthanna governorate. Males were outnumbering females. The highest age group affected was 0-10 years. The majority of patients were from rural areas. The majority of the cases presented within two months, involved the face, had a single lesion, and were of a dry type.

First Report of Natural Infection of Phlebotomus mongolensis to Leishmania major and Leishmania turanica in the Endemic Foci of Zoonotic Cutaneous Leishmaniasis in Iran.

The primary aim of this study is to determine infection to Leishmania parasites in the wild population of Phlebotomus caucasicus and Phlebotomus mongolensis using molecular methods in some important zoonotic cutaneous leishmaniasis foci in Iran. Sand flies were collected from active colonies of rodent burrows from 16 trapping sites using sticky trap paper. In order to detect and identify of Leishmania parasites in females Ph. caucasicus and Ph. mongolensis, the Nested-PCR amplification of ITS2-rDNA region was performed to generate amplicon with 245bp for Leishmania major, 206bp for L. gerbilli and 141bp for L. turanica. In the current study we found DNA of different gerbil parasites such as L. major and L. turanica, and mixed infection of L. major/L. turanica in Ph. caucasicus and Ph. mongolensis. It should be noted that, in Iran, natural infection with Leishmania parasites is recorded for the first time in this study in Ph. mongolensis. Both species of Ph. caucasicus and Ph. mongolensis not only may participate in the ZCL transmission cycle between reservoir hosts, but also results of this study support the role of these species as secondary vectors in the transmission of leishmaniasis to humans.

Molecular Detection of Leishmania Infection in Phlebotomine Sand Flies from an Endemic Focus of Zoonotic Cutaneous Leishmaniasis in Iran.

Due to the outbreak of zoonotic cutaneous leishmaniasis (ZCL), a disease caused by Leishmania major and mainly transmitted by Phlebotomus papatasi, in Damghan City, Semnan Province, the probable vectors of the disease were investigated in the city from 20 March 2016 to 20 January 2018. Sand flies were collected from indoors and outdoors biweekly by sticky traps in different parts of the city. The trapped sand flies were stored in 70% ethanol. They were identified and checked for Leishmania infections using nested-PCR method and specific primers; CSB1XR, CSB2XF, LiR, and 13Z. Overall, 1862 phlebotomine sand flies of Ph. papatasi (48.8%), Ph. andrejevi (8.3%), Ph. caucasicus (7.7), Ph. mongolensis (2%), Ph. sergenti (1.2%), Ph. alexandri (0.7%), Sergentomyia murgabiensis sintoni (29.3%), and Se. sumbarica (2%) were collected indoors (31.1%) and outdoors (68.9%). The highest and lowest numbers of collected sand flies were belonging to Ph. papatasi (48.8%) and Ph. alexandri (0.7%) respectively. 2.2% of the examined sand flies were shown to be infected with L. major and all were belonging to Ph. papatasi. This study confirms the report of Ph. papatasi infection with L. major and also the existence of Ph. sergenti and Ph. alexandri, the potential vectors of L. tropica and L. infantum respectively, in Damghan City. According to the findings, it is necessary for health officials to plan and take action to prevent the occurrence of ZCL epidemic in the city as well as the occurrence of other forms of leishmaniasis.

The Therapeutic Effects of Curcumin-coated Gold Nanoparticle Against Leishmania Major Causative Agent of Zoonotic Cutaneous Leishmaniasis (ZCL): An In Vitro and In Vivo Study

We synthesized and characterized curcumin-coated gold nanoparticles (Cur@AuNPs) and investigated their stability, cytotoxicity, leishmanicidal activity in in vitro and in in vivo experiments. Cur@AuNPs synthesized through a simple one-pot green chemistry technique. The in vitro leishmanicidal activity of curcumin-coated gold nanoparticles against extracellular promastigotes and intracellular amastigotes of protozoan parasite Leishmania major (L. major) was determined by applying the tetrazolium reduction colorimetric quantitative MTT technique. For in vivo assessment, the footpad lesion size and parasite burden in two infection site organs including lymph nodes and footpads of susceptible BALB/c mice infected with L. major were measured. Mice immune responses in all study groups were quantified by measuring the levels of gamma interferon (IFN-γ) and interleukin-4 (IL-4). Viability of Leishmania promastigotes significantly diminished with the inhibition in promastigotes growth (IC50) of 64.79μg/mL and 29.89μg/mL for 24h and 48h, respectively. In vitro nanoparticles treatment efficiently cleared the L. major amastigotes explanted in macrophages but had no harmful toxicity on the mice cells. In the in vivo condition, in the treated infected BALB/c mice the CL lesion size, Leishmania parasite burden, and IL-4 were decreased, while IFN-γ was significantly increased. The results suggest that Cur@AuNP was an effective compound against Leishmania parasite in vitro and in vivo, efficiently induced T-helper 1 (Th1) responses and augmented host cellular immune responses, and ending in a reduced Leishmania parasite burden. Therefore, it may be identified as a novel potential therapeutic approach for the local therapy of zoonotic CL treatment with high cure rates.

Calomyscid Rodents (Rodentia: Calomyscidae) as a Potential Reservoir of Zoonotic Cutaneous Leishmaniasis in a Mountainous Residential Area in the Plateau of Iran: Inferring from Molecular Data of kDNA and ITS2 Genes of Leishmania Major.

Cutaneous leishmaniasis (CL), a neglected tropical disease, is an important health problem in Fars Province, southern Iran. Fars, the fourth most populous Province in Iran, is the center of both anthroponotic and zoonotic cutaneous leishmaniasis (ZCL). Rodents, the reservoir of Leishmania major, play an important role in transmitting ZCL. In the present study, we report Leishmania infection in calomyscid rodents for the first time in mountainous residential areas of Shiraz, the capital of Fars Province, in southern Iran. Rodents were trapped in urban mountainous areas. The skin, liver, and spleen of rodents were examined microscopically for Leishmania infection. In addition, DNA was extracted from the tissues and they were evaluated for Leishmania infection by targeting the kDNA and subsequent sequencing of the nuclear rDNA internal transcribed spacer two (ITS2) region. DNA of L. major was detected in the spleen and liver of calomyscid rodents. Molecular evolution based on DNA-sequencing of the ITS2 gene confirmed the taxonomic situation of the parasite as L. major. Our findings suggest the eco-epidemiological importance of calomyscid rodents in the foci of leishmaniasis in the mountainous residential area on the plateau of Iran. These rodents may play a role in the transmission of leishmaniasis in a residential area and could be considered a potential reservoir for CL.