Anthroponotic Cutaneous Leishmaniasis Research Articles

Genetic diversity in Leishmania infantum and Leishmania tropica isolates from human and canine hosts in northern Morocco

This study investigated nine provinces in northern Morocco and collected 275 skin scraping, 22 bone marrow aspirates, and 89 fine needle aspirations from suspected cutaneous leishmaniasis (CL) and visceral leishmaniasis (VL) patients and potentially infected dogs. Molecular analysis using ITS1 RFLP PCR and RT-PCR revealed a higher prevalence of L. infantum (66.18 %; χ2 = 28.804; df = 1; P-value = 8.01e−08) than L. tropica in skin scraping, with L. infantum being the sole causative agent for both VL and canine leishmaniasis. L. infantum was predominantly found in most provinces, while L. tropica was relatively more dominant in Taza Province. Discriminant Analysis of Principal Components (DAPC) revealed distinct clustering between L. tropica and the other three species. However, no small subset of SNPs could clearly differentiate between Infantum_CL, Infantum_VL, and CanL, as they likely share a significant genetic background. The high rate of L. infantum could be attributed to the abundance of sand fly species transmitting VL. In Taza Province, Phlebotomus sergenti, responsible for anthroponotic CL, is the most abundant species. DNA sequencing demonstrated sequence heterogeneity in L. infantum (variants 1 to 9) and L. tropica (variants 1 to 7). Phylogenetic analysis showed a distinct separation between L. tropica and L. infantum strains, with an overlap among L. infantum strains isolated from cutaneous, visceral, and canine cases, and dogs serving as the central population for L. infantum.

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.

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.

Molecular identification of Leishmania tropica in mammals occurring in human-inhabited areas of a cutaneous leishmaniasis endemic focus in North-West Pakistan.

Cutaneous Leishmaniasis is endemic in the tribal district of Khyber near the Pak-Afghan border and is caused by Leishmania tropica. In Pakistan, cutaneous leishmaniasis caused by L. tropica is considered anthroponotic and is thought to be maintained by a human-sand fly-human transmission cycle. Along with humans, other mammals may also be acting as reservoir hosts of leishmaniasis in the study area. To investigate the role of non-human mammals in the transmission of leishmaniasis, blood samples were collected from 245 animals from the CL endemic district of Khyber, Pakistan. Leishmania parasite in these samples was detected by amplifying the species-specific sequences in minicircle kinetoplast DNA, using PCR. L. tropica DNA was detected in 18 (7.35%) samples, comprising 11 cows (Bos taurus), 6 goats (Capra hircus), and 1 dog (Canus lupus familiaris). Only a single cow and dog had a leishmaniasis-like lesion, and the remaining positive samples were asymptomatic. None of the tested sheep (Ovis aries) and rat (Rattus rattus, Rattus norvegicus) was positive. The present study reports the first instance of molecular detection of L. tropica in domestic animals. Our study indicates that along with humans' cows, goats and dogs may also be playing an important role in the transmission of anthroponotic cutaneous leishmaniasis in district Khyber in particular and Pakistan in general.

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.

Descriptive and Geometric Morphometry of the Wings of Phlebotomus sergenti Populations in Central Morocco.

Phlebotomus sergenti, the proven vector of Leishmania tropica, the causative agent of anthroponotic cutaneous leishmaniasis, is widely distributed in Morocco. Previous works using molecular markers (Internal Transcribed Spacer 2 rDNA and Cytochrome B mtDNA) hypothesized the existence of multiple closely related populations of sand fly species (cryptic species) that would exhibit distinct vectorial capacities. This work studies morphotypic diversity using traditional and geometric morphometry analyses carried out on Ph. sergenti's wings from central Morocco, where active L. tropica transmission occurs for 30 years. Descriptive characteristics (size and shape) of the right wings were measured in Ph. sergenti's specimens collected from fourteen stations in central Morocco. Both traditional and geometric morphometry methods were used to analyse geographic variations in Ph. sergenti wing's size and shape. These analyses support the existence of distinct Ph. sergenti populations, enlightening significant phenotypic variations of Ph. sergenti's wings, regarding their size and shape, depending on geographic origin. In addition, traditional and geometric morphometric analyses of the wing's length, centroid size, β, ɵ, and γ distances allowed clear discrimination of Ph. sergenti sub-populations. These data pinpoint the adaptative ability of Ph. sergenti to local environmental conditions. Additional studies are now required to further shed light on the genetic structure of Ph. sergenti populations in Morocco.

Treatment of Cutaneous Leishmaniasis with Allopurinol Plus Itraconazole in Iran.

Cutaneous leishmaniasis (CL) is a serious tropical disease and a neglected health challenge in Iran. Although limited data are available regarding anthroponotic CL, cases resistant to meglumine antimoniate (Glucantime) are increasingly being reported. Via an open-label noncontrolled case series, allopurinol (10 mg/kg/day) plus itraconazole (3-4 mg/kg/day) were orally administered for 1 month to 27 patients (56 lesions) with anthroponotic CL, most of whom were resistant to Glucantime. A mean lesion size of 3.5 ± 1.9 cm at baseline was reduced to 0.6 ± 1.0 after 1 month of treatment. Excellent treatment response was observed in 85.7% of lesions after 1 month. Recurrence only occurred in one patient in the 3-month follow-up session. This study presents preliminary evidence that oral allopurinol plus itraconazole could be an effective treatment in patients with anthroponotic CL.

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.

Mathematical Analysis of an Anthroponotic Cutaneous Leishmaniasis Model with Asymptomatic Infection

This work aims mainly to study the impact of experiencing asymptomatic anthroponotic cutaneous leishmaniasis (ACL) infection on the overall dynamics and outcomes of the disease. Therefore, a deterministic model for the transmission dynamics of ACL of type SEAIS in the human host and SI in sandfly populations is proposed and mathematically analyzed. The model is shown to be well-posed. Its equilibrium and stability analyses are shown. The equilibrium analysis shows that the model has an ACL-free equilibrium that is proven to be locally and globally asymptotically stable if and only if R0&lt;1. In addition, the model has a unique ACL-endemic equilibrium that is shown to exist and be locally asymptotically stable if and only if R0&gt;1. Numerical simulations are performed to show the asymptotic stability of these equilibriums. In addition, the effect of ignoring asymptomatic infections is studied and the analysis shows that ignoring the development of asymptomatic infections overestimates the effort required to eliminate the infection. Moreover, it implies inaccurate measures of controlling ACL infection, especially those based on either using insecticide sprays or bednets.

The Potential Role of Lipopeptide Biosurfactant Generated by Acinetobacter junii B6 on Leishmania Tropica: The Synergy of Lipopeptide Biosurfactant and Glucantime

An in vitro investigation was carried out to assess how the lipopeptide biosurfactant (LPB) produced by Acinetobacter junii B6 affects Leishmania tropica infection and the associated cytokine gene expression in macrophages infected with L. tropica. Glucantime® (meglumine antimoniate, MA) and LPB were investigated for their leishmanicidal effect, alone and in combination (LPB + MA), using a colorimetric test and a macrophage model. Immunomodulatory impact was also evaluated through analysis of Th1 and Th2 cytokine gene expression in infected macrophages after treatment with MA and LPB, individually and in combination. The MA/LPB combination showed higher inhibitory impacts on L. tropica amastigotes and promastigotes than each alone. Cytokine gene expression confirmed LPB’s affinity to IFN-γ, affirming the elevated IL-12p40 and IFN- γ concentrations in addition to a reduction in the secretion of IL-10 in a dose-dependent manner, particularly in combined treatment. The results indicated higher effectiveness of LPB along with MA in the reduction of the parasite growth and promoting the immune reaction level, which may be considered as a possible therapeutic strategy to treat those with anthroponotic cutaneous leishmaniasis.

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.

Population Genetic Structure of Phlebotomus sergenti (Diptera: Psychodidae) Collected in Four Regions of Morocco Based on the Analysis of Cyt b and EF-1α Genes.

Phlebotomus (Ph.) sergenti is the main vector of Leishmania (L.) tropica (Trypanosomatida: Trypanosomatidae), the causative agent of anthroponotic cutaneous leishmaniasis in Morocco. This species has an extended geographical distribution, wider than that of the parasite. The main objective of our study was to analyze the genetic diversity of Ph. sergenti collected in four foci in Morocco: Taza, Foum Jemâa, El Hanchane, and Ouarzazate. We studied a set of diversity and population structure indices by sequencing two markers; nuclear EF-1α and mitochondrial Cyt b from 175 individual sand flies. Our results showed a considerable degree of intraspecific polymorphism with a high number of haplotypes identified in both genes. Many polymorphic sites detected in the Cyt b sequences (SCyt b = 45) indicate that it is the most polymorphic marker showing a distinct distribution of haplotypes according to their geographical origin, whereas the EF-1α marker showed no geographical isolation. Analysis by Tajima's D and Fu's Fs tests revealed a possible recent expansion of the populations, especially with the EF-1α marker, showing significant values in Taza and Ouarzazate sequences. The present study revealed significant genetic diversity within Ph. sergenti populations in Morocco. The results warrant further research using a combination of more than two markers including mitochondrial and non-mitochondrial markers, which may provide more information to clarify the genetic status of Ph. sergenti.

Overexpression of Iron Super Oxide Dismutases A/B Genes Are Associated with Antimony Resistance of Leishmania tropica Clinical Isolates.

Pentavalent antimonial has been a drug of choice against leishmaniasis, despite the emergence of treatment failure. Identification of resistance markers is urgently needed to design new therapeutic strategies. Iron-Superoxide dismutases (Fe-SODs) are antioxidant enzymes contributing to detoxify reactive oxygen species to prevent a cell from oxidative stress. Since antimonial compounds induce oxidative stress, in this survey, the expression of SOD genes was investigated to identify their expression pattern in clinical resistant isolates. This cross-sectional survey was done in Mashhad City, northeast of Iran during 2014 to 2019. The RNA expression level of mitochondrial (SODA) and glycosomal (SODB) superoxide dismutase was investigated in 25 antimony responsive (n=15) and unresponsive (n=10) anthroponotic cutaneous leishmaniasis (ACL) patients. Total RNA extraction and cDNA synthesis, the qRT-PCR approach was utilized to investigate the relative RNA expression level. The transcript level of SODs was over-expressed in the most resistant isolates. Gene expression analysis demonstrated the over-expression of SODA and B by a factor of 3.8 and 4.81, respectively, in resistance isolates vs. sensitive ones. Aberrant expression of SODA/B in unresponsive parasites could potentially implicate in detoxifying antimony-induced oxidative stress. Moreover, SODs might be considered as potential predictive markers of the response to antimonials in ACL patients in endemic areas.

Poor adherence is a major barrier to the proper treatment of cutaneous leishmaniasis: A case-control field assessment in Iran

Leishmaniasis is an overlooked, poverty-stricken, and complex disease with growing social and public health problems. In general, leishmaniasis is a curable disease; however, there is an expansion of unresponsive cases to treatment in cutaneous leishmaniasis (CL). One of the effective and ignored determinants in the treatment outcome of CL is poor treatment adherence (PTA). PTA is an overlooked and widespread phenomenon to proper Leishmania treatment. This study aimed to explore the effect of poor adherence in unresponsiveness to treatment in patients with anthroponotic CL (ACL) by comparing conventional statistical modalities and machine learning analyses in Iran. Overall, 190 cases consisting of 50 unresponsive patients (case group), and 140 responsive patients (control group) with ACL were randomly selected. The data collecting form that included 25 queries (Q) was recorded for each case and analyzed by R software and genetic algorithm (GA) approaches. Complex treatment regimens (Q11), cultural and lay views about the disease and therapy (Q8), life stress, hopelessness and negative feelings (Q22), adverse effects of treatment (Q13), and long duration of the lesion (Q12) were the most prevalent significant variables that inhibited effective treatment adherence by the two methods, in decreasing order of significance. In the inherent algorithm approach, similar to the statistical approach, the most significant feature was complex treatment regimens (Q11). Providing essential knowledge about ACL and treatment of patients with chronic diseases and patients with misconceptions about chemical drugs are important issues directly related to the disease's unresponsiveness. Furthermore, early detection of patients to prevent the long duration of the disease and the process of treatment, efforts to minimize side effects of treatment, induction of positive thinking, and giving hope to patients with stress and anxiety by medical staff, and family can help patients adhere to the treatment.

Determination of the feeding behavior of Phlebotomus sergenti using multiplex PCR and tent-baited traps in a new focus of Anthroponotic Cutaneous Leishmaniasis in the southeast of Iran

ObjectivesThis study aimed to determine feeding behaviors of Phlebotomus sergenti Parrot, in a new focus of Anthroponotic Cutaneous Leishmaniasis in Bam County, southeast Iran. MethodsTwo methods were used to determine the feeding behavior of Phlebotomus sergenti. In the first method, blood-fed sand flies were captured using a mouth aspirator in human and animal dwellings and consequently, blood meal identification was made using Multiplex PCR. The results were used for calculating Host Feeding Index (HFI) and Forage Ratio (FR) parameters. In the second method, human (Homo sapiens), goat (Capra aegagrus), cattle (Bos taurus), chicken (Gallus gallus) and dog (Canis lupus) were used as baits in tent-baited traps to determine the feeding behavior of Phlebotomus sergenti. ResultsMultiplex PCR analysis revealed that the most frequent blood in the stomack of sand flies’ were from chicken, but the calculation of the FR revealed that this species prefers canine and poultary blood as meal. Human and animal tent-baited traps revealed that most Phlebotomus sergenti were attracted to chicken rather than the other hosts. ConclusionsSand flies are attracted to animals for various reasons such as eating blood, mating on their bodies and laying eggs on their feces. Molecular methods are effective and accurate methods to determine the type of host that sandfly fed on, but they do not show host preferences. The results of the molecular analysis, along with the calculation of HFI and FR, can determine the preferred host of sand flies. The current study revealed that dogs, the secondary reservoir of ACL in Iran, is the first preferred host of Phlebotomus sergenti.

Determinants of anthroponotic cutaneous leishmaniasis by case-control study in Morocco.

Leishmaniasis is endemic in Morocco where both cutaneous and visceral forms coexist. To date, anthroponotic cutaneous leishmaniasis (ACL) determinants remain poorly investigated in Morocco. However, the disease risk factors identification is vital to determine the specific preventive process. In this aim, a case-control study was conducted in the main active ACL foci in central Morocco. Epidemiological data were extracted from bulletins, registers and annual reports of the regional direction of Health offices. The socioeconomic and environmental data were collected from epidemiological surveys, completed by a questionnaire intended for accessible positive population and control people selected from the cases' entourage. The study included 258 cases and 395 controls. Our results showed that many socioeconomic factors were associated with ACL in Morocco such as the rural habitation (OR = 4.163; 95% CI: 2.91-5.96), movement to endemic area (OR = 4.53; 95% CI: 3.03-6.77), provenance from leishmaniasis foci (in Essaouira focus OR = 5.34; 95% CI: 1.19-24.03) and poverty. In addition, environmental factors like proximity of vegetation (OR = 2.45; 95% CI: 1.14-5.25), poor domestic hygiene, particularly the absence of sewage system/waste management (OR = 1.63; 95% CI: 1.35-1.96), and presence of animals (OR = 2.67; 95% CI: 1.14-5.25) increase the risk of ACL in Morocco. Except for Matrimonial status (married people, OR = 4.11; 95% CI: 1.80-9.41), there is however no significant association of the disease with the other socio-demographic factors in the study area (p>0.05). These several risk factors must be taken in consideration to prevent this disease through multidisciplinary collaboration and community participation.

Patterns of cutaneous leishmaniasis during the COVID-19 pandemic in four endemic regions of Iran.

Both zoonotic cutaneous leishmaniasis (ZCL) and anthroponotic cutaneous leishmaniasis are endemic in different regions of Iran. It is important to know the type of cutaneous leishmaniasis (CL) for prevention strategies. On the other hand, Iran is one of the countries that was severely affected by the coronavirus 2019 (COVID-19) pandemic. However, little is known about the causative agents of CL during the COVID-19 pandemic in Iran. In this study, a total of 181 samples were isolated from patients with CL lesions in four different endemic cities of Iran (Jahrom, Juyom, Kashan and Shoushtar) during the COVID-19 pandemic (the summer and autumn of 2021). The nested-PCR targeting kinetoplast DNA (kDNA) minicircles was applied for species identification of the parasite and the internal transcribed spacer-1 (ITS-1) gene was applied for sequencing and phylogenetic analysis. Molecular detection of kDNA minicircles revealed that all isolates from the four cities were Leishmania major, indicating the ZCL pattern. Ten isolates were sequenced by the ITS-1 gene and deposited in GenBank [accession numbers: OL627363-72]. Sequencing and phylogenetic analysis displayed a high similarity rate of the isolates with sequences from other parts of Iran and Iraq. Over half of the patients (53.59%) had a single lesion, while 17.12, 14.92, 2.21 and 12.16% of cases had two, three, four and more than four lesions, respectively. About half of the patients (50.27%) had lesions on their hands, while the rest had lesions on multiple locations (19.34%), legs (16.58%), face (9.94%), ears (1.66%) and waist (2.21%). This study revealed the occurrence of ZCL during the COVID-19 pandemic in four endemic regions of Iran. Strategies for prevention and control of the disease should be considered to mitigate the occurrence of ZCL.

Climate Change Influences on the Potential Distribution of the Sand Fly Phlebotomus sergenti, Vector of Leishmania tropica in Morocco.

Leishmaniases are a vector-borne disease, re-emerging in several regions of the world posing a burden on public health. As other vector-borne diseases, climate change is a crucial factor affecting the evolution of leishmaniasis. In Morocco, anthroponotic cutaneous leishmaniasis (ACL) is widespread geographically as many foci across the country, mainly in central Morocco. The objective of this study is to evaluate the potential impacts of climate change on the distribution of ACL due to Leishmania tropica, and its corresponding vector Phlebotomus sergenti in Morocco. Using Ecological Niche Modeling (ENM) tool, the estimated geographical range shift of L. tropica and P. sergenti by 2050 was projected under two Representative's Concentration's Pathways (RCPs) to be 2.6 and RCP 8.5 respectively. P. sergenti records were obtained from field collections of the laboratory team and previously published entomological observations, while, epidemiological data forL. tropica were obtained from Moroccan Ministry of Health reports. Our models under present-day conditions indicated a probable expansion for L. tropica as well as for its vector in Morocco, P. sergenti. It showed a concentrated distribution in the west-central and northern area of Morocco. Future predictions anticipate expansion into areas not identified as suitable for P. sergenti under present conditions, particularly in northern and southeastern areas of Morocco. L. tropica is also expected to have high expansion in southern areas for the next 30years in Morocco. This indicates that L. tropica and P. sergenti will continue to find suitable climate conditions in the future. A higher abundance of P. sergenti may indeed result in a higher transmission risk of ACL. This information is essential in developing a control plan for ACL in Morocco. However, future investigations on L. tropica reservoirs are needed to confirm our predictions.

Involvement of tryparedoxin peroxidase (TryP) and trypanothione reductase (TryR) in antimony unresponsive of Leishmania tropica clinical isolates of Iran

Clinical resistance to pentavalent antimonial compounds has long been recognized as a major problem in the treatment of human leishmaniasis. Trypanothione metabolism, the main form of thiol, has shown to play a central role in antimony resistance of laboratory-generated resistant Leishmania spp. and field-isolated resistant L. donovani; but the mechanism of antimony resistance in the clinical isolates of L. tropica causing anthroponotic cutaneous leishmaniasis (ACL) is less studied. Patients were selected among confirmed positive ACL cases who referred to Pasteur Institute of Iran, Tehran, from endemic regions of north-east and south of Iran. L. tropica clinical isolates were collected from patients who were either treatment-responsive (MAS=S1 to S5) or unresponsive (MAR=R1 to R4) to Glucantime® (meglumine antimoniate=MA). Isolates were tested for sensitivity to trivalent antimony (SbIII) in promastigotes and to pentavalent antimony (SbV) in intracellular amastigotes stages. Intracellular thiol levels were assayed and trypanothione-dependent components, including trypanothione reductase (TR) and tryparedoxin peroxidase I (TryP) were analysed at protein level and enzymatic activity in isolates. The MAR isolates had an approximate two fold increase in the levels of intracellular thiols (P< 0.05) accompanied by an average 5-10 fold increase in in vitro resistance to antimony. TryP was amplified at the protein level in all MAR strains as compared to the MAS strains (range: 2.8-5.6 fold). All MAR isolates metabolized H2O2 at higher rates than MAS isolates (8.55±0.75 nmol/min/mg vs. 3.14±0.36 nmol/min/mg) (P< 0.05). In addition, levels of TryR protein were also markedly elevated in 3 out of 4 MAR isolates (range: 2.2-4.1 fold). This was accompanied by overexpressed TryR activity (mean level of 46.83±2.43 for extracts of MAR vs. 20.98±3.02 for MAS strains) (P< 0.05).Elevated levels of TryP, active enzyme in peroxide detoxification, were observed in MAR parasites resulting in an increased metabolism of H2O2. TryR activity was overexpressed on average in extracts of MAR strains, but not in all isolates. Enhanced anti-oxidant defenses through thiol metabolism may play a significant role in clinical resistance of ACL patients to Glucantime.

Combination of topical liposomal amphotericin B and Glucantime in comparison with glucantime alone for the treatment of anthroponotic cutaneous leishmaniasis (ACL) caused by Leishmania tropica: study protocol for a randomized, controlled trial.

Background and Objectives:Cutaneous leishmaniasis (CL) treatment is a challenging issue, although numerous modalities have been introduced as candidate treatment for CL yet only antimonial agents are commonly used to treat CL, a different form of amphotericin B is used to treat visceral form of leishmaniasis but the efficacy against CL is not high. There are a few reliable clinical trials on CL, the main reason is the nature of the disease which required a well design protocol to evaluate the efficacy of any candidate treatment against CL. In this study, a protocol was developed and used to evaluate a topical formulation of a nano-liposomal form of amphotericin B in addition to glucantime to treat CL caused by L. tropica.Materials and Methods:This study is a phase 3, randomized, double-blind, placebo-controlled trial to evaluate the efficacy and safety of topical nano-liposomal amphotericin B (SinaAmpholeish 0.4%) in combination with intralesional injections of meglumine antimoniate in the treatment of ACL caused by L. tropica. Overall, 130 patients, aged 12-60 years, with a diagnosis of ACL caused by L. tropica are recruited and treated according to the protocol.Results:A total of 130 patients with CL lesion will be recruited and doubleblind randomly treated with received intralesional injections of Glucantime weekly or Glucantime plus SinaAmpholeish for 4 weeks.Conclusion:The results of this study showed that the protocol works well and the treatment was tolerated by both groups of patients.