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Background: Leishmaniasis is an important public health parasitic infection, which is endemic in many parts of the world, including Iran. We aimed to investigate genetic diversity and phylogenetic relationship among different Leishmania isolates using multilocus sequence typing (MLST). Methods: Totally, 41 isolates collected either from patients referred to Leishmaniasis Diagnostics and Treatment Center at Tehran University of Medical Sciences, Tehran, Iran or from animals during 2019-2021, were subjected to the study. They included L. major and L. tropica from human, L. infantum from canine, and L. turanica from rodents from different endemic foci of Iran analyzed using MLST including gp63, g6pdh, lack, nagt, and hsp70 genes. Results: A total of 5010 bps was analyzed from each isolate. The three targets, nagt, lack, and g6pdh, generated better topology comparing to the other genes. In the 44 isolates, 22 haplotypes (STs) were identified. Leishmania tropica contained the highest number of haplotypes (n=12) comparing to L. major (n=8), L. infantum (n=1) and L. turanica (n=1). All five genomic loci caused separation of Iranian Leishmania species at the species level, indicating conservation of these genes in the Leishmania parasite. Conclusion: The highest number of haplotypes belonged to L. tropica, indicating that the genetic diversity of this species is higher than that of L. major. It was further confirmed that the MLST is a suitable method to examine genetic variation of Leishmania parasites with respect to evolutionary and epidemiological studies. [ABSTRACT FROM AUTHOR]

Background: Recent studies have shown an increasing number of patients with cutaneous leishmaniasis (CL) who do not respond to pentavalent antimonials as the first line of treatment for CL. Nanocarriers such as extracellular vesicles (EVs) are efficient vehicles that might be used as drug delivery systems for the treatment of diseases. Therefore, we aimed to isolate and characterize the EVs of Leishmania major, load them with Amphotericin B (AmB), and investigate the toxicity and efficacy of the prepared drug form. Methods: The EVs of L. major were isolated, characterized, and loaded with amphotericin B (AmB), and the EVs-Amphotericin B (EVs-AmB) form was synthesized. Relevant in vitro and in vivo methods were performed to evaluate the toxicity and efficacy of EVs-AmB compared to the control. Results: The anti-leishmanial activity of the EVs-AmB showed a higher percentage inhibition (PI%) (P = 0.023) compared to the AmB at different concentrations and time points. Obtained data showed a significant increase in the lesion size and parasite load in the lesion, PBS, and EVs mice groups in comparison with EVs-AmB, AmB, and Glucantime groups (P < 0.05), EVs-AmB had a significant decrease in lesion sizes in comparison with AmB (P < 0.05). Results showed that EVs-AmB decreased its toxicity to the kidneys and liver (P < 0.05). Conclusion: EVs-AmB improved the efficacy of AmB in mouse skin lesions and reduced hepatorenal toxicity. Furthermore, EVs could be a promising nanoplatform for the delivery of AmB in CL caused by L. major. [ABSTRACT FROM AUTHOR]

Background: We aimed to verify the susceptibility of Leishmania infantum, L. major and L. tropica, to commercial lectins in order to identify the three Leishmania species. Methods: The degree of agglutination was determined both macroscopically and microscopically and was scored negative (-) to positive (from 1+- 4+) based on their percentage of agglutination. Results: Jacalin and UEA-1 were capable of agglutination of L. infantum isolates in both logarithmic and stationary phases at a concentration of 1000 µg/ml (100%). L. tropica isolates showed agglutination with the lectin UEA-1 in both logarithmic and stationary phases (62.5% and 87.5%). L. major and L. tropica showed 75% agglutination with lectin Jacalin in both logarithmic and stationary phases. L. tropica isolates showed 25% agglutination with the lectin WGA in the logarithmic phase. L. infantum, L. major and L. tropica isolates showed 25, 12.5 and 37.5% agglutination in the stationary phase, however, did not show agglutination in logarithmic phases. L. major isolates showed 12.5% agglutination with the lectin PHA in the stationary phase, however, were incapable of agglutination with the L. tropica and L. infantum in both logarithmic and stationary phases. Conclusion: Despite the fact, that JCA and I-UEA lectins were not able to completely separate L. infantum, L. major and L. tropica. WGA lectin and PHA lectin can help in separating the species of Leishmania parasites. [ABSTRACT FROM AUTHOR]

Background: Visceral leishmaniasis (VL) is one of the most important neglected tropical diseases. The zoonotic form of VL is endemic in some areas of Iran. We aimed to determine the status of VL identified in humans and canines in different parts of Iran from 2013 to 2022. Method: A national representative cross-sectional study was conducted in 10 provinces of Iran, including the national leishmaniasis reference lab. We employed the direct agglutination test (DAT) as a reliable serological method to detect anti-Leishmania infantum antibodies in humans and animal reservoir hosts. Additionally, a narrative literature review was conducted to identify relevant studies on VL seroprevalence in Iran from 2013 to 2023. Results: The results of 21281 human and 5610 canine serum samples from 2013 to 2022 are reported. Altogether, 448 (2.1%, 95%CI: 2.0-2.3) human serum samples showed anti-L. infantum antibody levels of ≥1:3200. Of these samples, 13716 (64.5%) were collected actively, which showed a seroprevalence of 0.6% (95% CI: 0.5-0.8) and 7565 (35.5%) were collected passively, which showed a seroprevalence of 4.8% (95%CI: 4.3-5.3). Overall, 1035 (20.1%, 95%CI: 19.0-21.2) of 5160 domestic dogs (Canis familiaris) samples showed anti-L. infantum antibody levels of ≥1:320. Northwest (...,^%) and northeast (0.96%) regions had the highest human VL seroprevalence, while northwest (21.5%) and south (14.4%) regions had the highest canine VL seroprevalence. Conclusion: Zoonotic VL, an endemic parasitic disease, is still present in several different distinct areas across Iran. While human VL cases have shown a declining trend over the last decade, the prevalence of canine VL remains significant. [ABSTRACT FROM AUTHOR]

Background: Leishmaniasis is currently considered a re-emerging or emerging infection based on the geographic region. The outcome of leishmaniasis vastly depends on Leishmaniahost interaction. This preliminary study aimed to show the association of human leukocyte antigen (HLA) class I and II genes with healed and non-healed cutaneous leishmaniasis (CL), and symptomatic and asymptomatic visceral leishmaniasis (VL) compared with control groups in Iran. Methods: Ninety-five people, including 31 patients versus 64 individuals in the control group, were enrolled. Among them, 20 patients had confirmed CL based on amastigote observation, 10 had improved CL and 10 non-healed CL. Eleven patients were suffering from confirmed VL based on direct agglutination test (Five asymptomatic and six symptomatic VL cases). Besides, they were residents in an endemic area of VL in the northwest of Iran. To select a control group, it was ensured that they had no history of leishmaniasis. Peripheral blood samples were collected from each patient. After DNA extraction, HLA typing was conducted using polymerase chain reaction - sequence-specific priming (PCR-SSP). Subsequently, data were statistically analyzed by SPSS. Results: There was a statistical relationship between the presence of HLA-A26 and CL, healed CL and the existence of the B38 allele, C1 allele and symptomatic VL, as well as B1.4 allele and asymptomatic VL (P<0.05). Conclusion: This primary finding indicates that several HLA genes have a potential role in the susceptibility of Iranian people to CL and VL. [ABSTRACT FROM AUTHOR]

Background: Several markers have been described to characterise the population structure and genetic diversity of Fasciola species (Fasciola hepatica(F. hepatica) and Fasciola gigantica (F. gigantica). However, sequence analysis of a single genomic locus cannot provide sufficient resolution for the genetic diversity of the Fasciola parasite whose genomes are ∼1.3 GB in size. Objectives: To gain a better understanding of the gene diversity of Fasciola isolates from western Iran and to identify the most informative markers as candidates for epidemiological studies, five housekeeping genes were evaluated using a multilocus sequence typing (MLST) approach. Methods: MLST analysis was developed based on five genes (ND1, Pepck, Pold, Cyt b and HSP70) after genomic DNA extraction, amplification and sequencing. Nucleotide diversity and phylogeny analysis were conducted on both concatenated MLST loci and each individual locus. A median joining haplotype network was created to examine the haplotypes relationship among Fasciola isolates. Results: Thirty‐three Fasciola isolates (19 F. hepatica and 14 F. gigantica) were included in the study. A total of 2971 bp was analysed for each isolate and 31 sequence types (STs) were identified among the 33 isolates (19 for F. hepatica and 14 for F. gigantica isolates). The STs produced 44 and 42 polymorphic sites and 17 and 14 haplotypes for F. hepatica and F. gigantica, respectively. Haplotype diversity was 0.982 ± 0.026 and 1.000 ± 0.027 and nucleotide diversity was 0.00200 and 0.00353 ± 0.00088 for F. hepatica and F. gigantica, respectively. There was a high degree of genetic diversity with a Simpson's index of diversity of 0.98 and 1 for F. hepatica and F. gigantica, respectively. While HSP70 and Pold haplotypes from Fasciola species were separated by one to three mutational steps, the haplotype networks of ND1 and Cyt b were more complex and numerous mutational steps were found, likely due to recombination. Conclusions: Although HSP70 and Pold genes from F. gigantica were invariant over the entire region of sequence coverage, MLST was useful for investigating the phylogenetic relationship of Fasciola species. The present study also provided insight into markers more suitable for phylogenetic studies and the genetic structure of Fasciola parasites. [ABSTRACT FROM AUTHOR]

Background: 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. Methods: 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. Results: 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. Conclusion: 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. [ABSTRACT FROM AUTHOR]

Background: Visceral leishmaniasis (VL) also known as Kala-azar is considered as one of the zoonotic infections in Mediterranean countries. The disease reservoir and vectors are dogs and sandflies respectively. Due to reported sporadic cases of Kala-azar in the past five years in Shahroud County, Semnan Province, Iran, this study aimed to investigate the status of this infection in this area and to determine its seroepidemiology to take required measurements for infection control and treatment. Methods: This study was conducted on 504 subjects residing in seven villages in Shahroud County. Blood samples were randomly selected using the cluster sampling method and were collected from subjects aged up to 13 years old (90%) and adults over 13 years old (10%) from September to May 2019. After separating sera from whole blood, samples were subjected to direct agglutination test (DAT) to detect anti-Leishmania infantum antibodies. A range of 1:10 to 1:800 dilutions were prepared from the samples. Results: Results of 1:800 titration indicated that no sample was positive for antibodies against L. infantum. After the secondary screening, 10 cases (1.98%) showed the antibody titer of 1:100, while four cases (0.79%) showed the antibody titer of 1:400. Of 14 cases with the L. infantum antibodies, all were detected from the children <13 years old. According to clinical findings, no patient was suffering from fever, weight loss, splenomegaly, hepatomegaly, and cachexia and therefore did not show the Kala-azar symptoms. Conclusion: The results of the current study indicate that Kala-azar is not prevalent in Shahroud County. [ABSTRACT FROM AUTHOR]

Background: Detection of Leishmania RNA virus (LRV) in Old World Leishmania species and their possible role in the disease prognosis requires sensitive and specific methods, preferably independent of the viral genome. We aimed to develop an indirect immunofluorescence antibody (IFA) assay to detect LRV in the Old World Leishmania parasites. Methods: Clinical samples were collected from 86 cutaneous leishmaniasis (CL) patients in different endemic areas of CL in Iran, during 2017-2019. For antibody preparation, the viruses were obtained from sediment of an LRV-infected L. major culture-using freeze and thaw cycles followed by gradient cesium chloride centrifugation. The purified viruses were used to immunize a male 3-4 months rabbit. Various dilutions of the LRV-immunized rabbit's serum and a conjugated antibody were deployed to detect LRV in 48 isolates by IFA assay. Results: LRV virus was detected in four of the 48 CL cases using IFA method. Amplification of a partial fragment of RNA-dependent RNA polymerase (RdRp) gene from the isolates confirmed the IFA results. In phylogeny, the generated RdRp sequences from four isolates were grouped with the other Old World LRVs, but separate from L. aethiopica LRVs, which appeared as a highly supported distinct clade. Conclusion: Further optimization of this approach to detect the LRV directly in lesion scrapings can make it a more reliable tool for field studies and disclosing the virus's possible role in disseminating and unusual clinical features. [ABSTRACT FROM AUTHOR]

Background: The use of antimalarial drugs with number of compounds in combination form may potentiate each other's activity. Methods: This study was conducted in the School of Public Health, Tehran University of Medical Sciences, Tehran, Iran in 2018. It was based on two methods including in vivo and in vitro tests with aim of considering interaction between chitosan and chloroquine against Plasmodium berghei and P. falciparum parasites using different ratios of the agents with ED50s and IC50s baselines. Results: Administrating 10 and 20 mg/kg (mouse body weight) of chitosan alone to the P. berghei --infected mice up to 4 successive days resulted in 37% and 45% inhibition of P. berghei respectively, while employing the compound with chloroquine in combination form with ratios of 90/10 and 70/30 (chloroquine/chitosan) had a considerable potentiation including 71.58% and 83.85% inhibition effectiveness against P. berghei. Moreover, 20 mg/L (CCM) concentration of chitosan alone could eliminate 69.55% of P. falciparum in culture medium while in combination with chloroquine in ratios of 90/10 (chloroquine/chitosan) had considerable potentiation including 79.14% inhibition effectiveness. Mean survival time of those mice received combination therapy in ratios of 90/10 and 70/30 (chloroquine/chitosan) was longer than those took up mono therapy of either chloroquine or chitosan based on their ED50s doses. Conclusion: Interaction between chloroquine and chitosan showed considerable potentiation in combination form against either P. berghei or P. falciparum using in vivo and in vitro tests respectively. Meanwhile, interaction between the above mentioned agents resulted in a notable survival time for those P. berghei-infected mice treated with the combination. [ABSTRACT FROM AUTHOR]

Background: Cutaneous leishmaniasis (CL) is a vector borne disease predominantly found in tropical and subtropical countries, including Iran. For more than 6 decades, pentavalent antimonials have been used successfully worldwide for the treatment of leishmaniasis, but over the past few years, clinical resistance to these medications has increased. In this study, we evaluated CL patients who did not show any desirable responses to the anti-leishmanial treatment within a 10- year period (2008 to 2017). Methods: All patients from different parts of Iran suspected of having cutaneous leishmaniasis, who were referred to the laboratory of leishmaniosis in Tehran University of Medical Sciences from 2008–2017 were parasitological examined. Results: During this period, a total of 1480 suspected CL patients were referred to the laboratory of leishmaniosis. Samples from 655 patients (70.8%) suspected of having CL were positive microscopically. The failure rate in patients treated with anti-leishmaniasis medications for a minimum of three complete treatment periods was 1.83% (12 cases). There was no association between the number and size of skin lesions and patient characteristics. Also, the route of drug administration had no significant effect on the number and size of lesions. Conclusion: In the present study, treatment failure was found in some confirmed CL patients treated with meglumine antimoniate. Over the past few years, it seems that had been increased in resistance to these medications. So, a review of the correct implementation of the treatment protocol and/or a combination therapy may be helpful in preventing an increase in the rate of treatment failure. [ABSTRACT FROM AUTHOR]

Background: Mediterranean form of visceral leishmaniasis (VL) is endemic among some provinces of Iran. The present study was designed to determine the prevalence of canine visceral leishmaniasis (CVL) in the owned dogs of the rural areas of Alborz Province near Tehran as the capital of Iran. Methods: This study conducted on 303 owned dogs that selected using a stratified random sampling method. The direct agglutination test (DAT) was used to determine the frequency of Vl. The spleen biopsy was taken from the serologypositive dogs for the confirmation of CVL in the suspected dogs. Nested PCR and sequencing methods were used to determine the type of Leishmania species in the dogs which were parasitological positive. Results: Overall, the DAT results of 9 dogs (2.97%, CI: 1.57-5.55) showed anti Leishmania antibodies at titers ≥ 1:320 indicating VL infection. One dog (0.33%, CI 95%: 0.06-1.85) showed clinical signs and symptoms of VL. There was a significant correlation between the positive cases of CVL and rural area (p< 0.001). The Leishmania was observed in the impression smears that were prepared from spleen biopsy of five the studied dogs. Leishmania infantum were confirmed in all them using nested-PCR assay. The sequence analysis of all five isolates was 95% similar to L. infantum. Conclusion: This study shows that domestic cycle of L. infantum has been established in rural areas of Alborz province where located near Tehran as capital city of Iran. It is necessary to increase the awareness and monitoring of the disease periodically. [ABSTRACT FROM AUTHOR]

Background: Visceral leishmaniasis (VL) is the most severe form of leishmaniasis in Iran with high mortality rates in the case of inaccurate diagnosis and treatment. This study aimed to prepare and evaluate a new rk39 recombinant antigen from an Iranian strain of Leishmania infantum for diagnosis of VL in humans and dogs. Methods: rK39-based enzyme-linked immunosorbent assay (ELISA) was compared with the direct agglutination test (DAT) for the detection of anti L. infantum antibodies. We screened 84 human sera and 87 dog sera from clinical cases in the endemic area of Meshkin-Shahr, Iran along with 176 sera from healthy controls (collected from 86 humans and 90 dogs) during 2013 -2016. Results: Using the rK39 ELISA, a sensitivity of 85.7% (95% CI, 95-99%) and a specificity of 86.0% (95% CI, 95%-99%) were detected in human sera at a 1:800 (cut-off) titer when DAT-confirmed cases were compared with healthy controls; a sensitivity of 96.6% (95% CI, 95%-99%) and specificity of 94.4% (95% CI, 95%-99%) were found at a 1:80 (cut-off) titer compared with DAT. Kappa analysis indicated agreement between the rK39 ELISA and DAT (0.718) when using human sera at a 1:800 (cut-off) titer as well as (0.910) at a 1:80 (cut-off) titer when using dog sera (P<0.05). Conclusion: New rk39 recombinant antigen from an Iranian strain of Leishmania infantum seems to be used for diagnosis of VL in humans and dogs. Further extended field studies are recommended. [ABSTRACT FROM AUTHOR]

Background: To overcome human malaria problem several solutions have been employed including extensive studies in the field of Plasmodia relevant antigens. The aim of this study was to determine allelic variation in the MSP1 gene of Plasmodium falciparum among some falciparum malaria-infected patients in Southeastern Iran. Methods: Twenty P. falciparum positive cases were enrolled from Sistan and Baluchistan Province, southeastern Iran in 2013-15. From each case, 1.5ml of peripheral blood was collected into EDTA contained tubes. Thick and thin blood smears were stained with standard Giemsa stain and were checked with conventional microscopical method. DNA was extracted from blood samples and amplification of block 2 MSP1 was performed using specific primers. Gel electrophoresis was done and results showed some amplification fragments corresponding to block 2 regions of Pf MSP1 gene. Finally, four samples from different allelic types were sent for sequencing process. Results: Fragments were different in size, so classified into six allelic types as kinds of 1-6 based on happening frequencies. Digestion of PCR products revealed two sub allelic types (A and B) within allelic types 2 and 3, but not in allelic types 1, 4, 5 and 6. Twenty percent of samples were sent for sequencing. Sequence alignment showed 78.95% to 91.83% identity between samples. Conclusion: Identity between samples and phylogenetic tree revealed that there is an extensive diversity range among isolates. Fifty percent of the isolates were under the risk of complicated malaria. Two of these patients (10%) needed special care and recovery was obtained after getting hospital services. [ABSTRACT FROM AUTHOR]

Background: Cutaneous leishmaniasis (CL) is described as a major health problem in many countries of the world. Regulatory T cells (Tregs) are characterized as one of immunologic indexes. One of the best methods to determine of Tregs percentage is flow cytometry. The aim of this study was determination of the role of Tregs profile among acute and chronic forms of human CL using flow cytometry analysis. Methods: This study was conducted on 24 patients referred to Laboratory of Leishmaniasis, Tehran University of Medical Sciences, Tehran, Iran with acute and 14 patients with chronic phases of CL as well as 15 healthy individuals as control group in 2015-2016. After microscopic examination, 2 ml of peripheral blood samples were collected for determining percentage of CD4 + CD25 + CD127 low Tregs by using flow cytometry method. Results: Using flow cytometry analysis, the average percentage of Tregs were calculated 5.73, 6.71 and 6.61 for acute, chronic and healthy individuals, respectively. With SPSS software and Scheffe multiple comparison tests, the differences within in these groups are statistically significant (P=0.04) and between the acute and chronic group, there was marginally significant with approximately 91% of confidence level (P=0.088). Conclusion: Marginally differences were found significantly among averages of Regulatory T cells, acute and chronic phases of CL. Further comprehensive studies can be needed to verify the role of Tregs in both phases of CL cases. [ABSTRACT FROM AUTHOR]

Background: Visceral leishmaniasis (VL) is endemic in the northwest and south of Iran. Untreated cases of VL could cause death. The aim of the present study was to evaluate the diagnostic performance of western blotting to detect a specific immunodominant proteins pattern for Leishmania infantum infection using human sera infected with VL. Methods: We studied a panel of 122 cryopreserved human serum samples from the leishmaniasis Research Laboratory, Tehran University of Medical Sciences, Tehran, Iran from 2010 to 2017. Serum samples were collected from visceral (Group I, n: 43) and cutaneous leishmaniasis (CL) (Group II, n: 8) patients, healthy individuals from endemic (Group III, n: 13) and non-endemic (Group IV, n: 16) areas for VL, and patients with other infectious diseases (Group V, n: 42). Total antigens were prepared from the Iranian strain of L. infantum promastigote form. Results: In western blotting method, 34 protein bands of 14 to 163 kDa were recognized using the sera of VL patients. The polypeptide fractions with the highest frequency including 29, 51, and 62 kDa fractions were detected using 81.4%, 79%, and 81.4% of the sera, respectively. These bands were not detected using the sera of the negative control. Moreover, 19–23, 27, 31–35, 143–163, and 109 kDa fractions were detected specifically using the sera of the patients with VL. Conclusion: This technique could be a primary step for further exploration of VL immunodominant antigens for cloning (or any technique) further investigations for future planning. [ABSTRACT FROM AUTHOR]

Background: Leishmaniasis is a major medical health problem and distributes in nearly half of 31 provinces of Iran. We aimed to identify cutaneous and visceral Leishmania spp. isolated from infected humans and domestic dogs in various regions of Iran, 2010-2013. Methods: DNA was extracted from 108 lesion exudate samples of suspected patients to cutaneous leishmaniasis and nine liver and spleen aspirates of infected dogs cultured in RPMI-1640 and amplified using partial sequence of ITS1 gene. The PCR amplicons were digested using HaeIII endonuclease enzyme and used in restriction fragment length polymorphism (RFLP) assay. Then, 48 amplicons representing various hosts were sequenced and compared to sequences from GenBank databases using BLAST. Results: PCR-RFLP analysis showed that 60 and 48 CL patients were infected by Leishmania tropica and L. major, respectively. From nine canine visceral leishmaniasis (CVL) isolates, eight isolates were identified as L. infantum and one as L. tropica. The greatest similarity of 95.7% in ITS1 region was seen between L. infantum and L. major. Furthermore, the lowest similarity with 65.7% was seen between L. tropica and L. major. Intra-species comparison of ITS1 region in L. infantum, L. major and L. tropica isolates were showed 100%, 98.2% and 72.4 % similarities, respectively. Conclusion: PCR-RFLP based on ITS1 region is an appropriate method to distinguish three Leishmania spp. of L. major, L. tropica, and L. infantum. In intra-species comparison of ITS1 region, genotypic variations showed that L. tropica isolates were more heterogeneous than L. major and L. infantum isolates. [ABSTRACT FROM AUTHOR]

Background: Cutaneous leishmaniasis (CL) is one of the world health problems. Leishmania major is the etiological agent of zoonotic cutaneous leishmaniasis (ZCL). Promastigote and amastigote are two morphological forms of Leishmania parasites that express different proteins and p27 is an important gene encoding cytochrome c oxidase (COX) component. P27 gene expresses a 27 kDa protein that essential in ATP synthesis. This study aimed to compare p27 gene expression in promastigote and amastigote forms in Iranian strain of L. major (MRHO/IR/75/ER). Methods: This study was conducted in 2015. Clinical isolates of CL patients from north, center, west and south parts of Iran were collected and identified by PCRRFLP. After RNA extraction of promastigotes and amastigotes and cDNA synthesis, the expression level of p27 gene was compared by real-time RT-PCR. Results: By comparison of expression level between amastigote and promastigote forms of Iranian strain of L. major, up-regulation of p27 gene (2.73 fold) was observed in amastigotes. Moreover, there was no significant difference in p27 gene expression between L. major isolates. Conclusion: p27 gene and protein can be considered as a target in recombinant vaccine production and treatment process. [ABSTRACT FROM AUTHOR]

Despite the broad distribution of leishmaniasis among Iranians and animals across the country, little is known about the genetic characteristics of the causative agents. Applying both HSP70 PCR-RFLP and sequence analyses, this study aimed to evaluate the genetic diversity and phylogenetic relationships among Leishmania spp. isolated from Iranian endemic foci and available reference strains. A total of 36 Leishmania isolates from almost all districts across the country were genetically analyzed for the HSP70 gene using both PCR-RFLP and sequence analysis. The original HSP70 gene sequences were aligned along with homologous Leishmania sequences retrieved from NCBI, and subjected to the phylogenetic analysis. Basic parameters of genetic diversity were also estimated. The HSP70 PCR-RFLP presented 3 different electrophoretic patterns, with no further intraspecific variation, corresponding to 3 Leishmania species available in the country, L. tropica, L. major, and L. infantum. Phylogenetic analyses presented 5 major clades, corresponding to 5 species complexes. Iranian lineages, including L. major, L. tropica, and L. infantum, were distributed among 3 complexes L. major, L. tropica, and L. donovani. However, within the L. major and L. donovani species complexes, the HSP70 phylogeny was not able to distinguish clearly between the L. major and L. turanica isolates, and between the L. infantum, L. donovani, and L. chagasi isolates, respectively. Our results indicated that both HSP70 PCR-RFLP and sequence analyses are medically applicable tools for identification of Leishmania species in Iranian patients. However, the reduced genetic diversity of the target gene makes it inevitable that its phylogeny only resolves the major groups, namely, the species complexes. [ABSTRACT FROM AUTHOR]

Background: Demodicosis is one of the most prevalent skin diseases resulting from infestation by Demodex mites. This parasite usually inhabits in follicular infundibulum or sebaceous duct and transmits through close contact with an infested host. Methods: This study was carried from September 2014 to January 2016 at Tehran University of Medical Sciences, Tehran, Iran. DNA extraction and amplification of 16S ribosomal RNA was performed on four isolates, already obtained from four different patients and identified morphologically though clearing with 10% Potassium hydroxide (KOH) and microscopical examination. Amplified fragments from the isolates were compared with GeneBank database and phylogenetic analysis was carried out using MEGA6 software. Results: A 390 bp fragment of 16S rDNA was obtained in all isolates and analysis of generated sequences showed high similarity with those submitted to GenBank, previously. Intra-species similarity and distance also showed 99.983% and 0.017, respectively, for the studied isolates. Multiple alignments of the isolates showed Single Nucleotide Polymorphisms (SNPs) in 16S rRNA fragment. Phylogenetic analysis revealed that all 4 isolates clustered with other D. folliculorum, recovered from GenBank database. Our accession numbers KF875587 and KF875589 showed more similarity together in comparison with two other studied isolates. Conclusion: Mitochondrial 16S rDNA is one of the most suitable molecular barcodes for identification D. folliculorum and this fragment can use for intra-species characterization of the most human-infected mites. [ABSTRACT FROM AUTHOR]

Background: Cutaneous leishmaniasis (CL) is one of the most important neglected tropical diseases and a major public health challenge in Iran caused by Leishmania spp and transmitted by phlebotomine sand flies. The number of CL cases has shown an increasing pattern all over the country, including the district of Varamin, southeast of Tehran, Iran. This study aimed to identify the Leishmania spp isolated from CL patients using molecular methods in Varamin during 2012-2013. Methods: Exudate materials collected from the swollen edge of the skin lesions of 44 parasitological positive CL patients by disposable lancet. They were referred to Varamin Health Center by physician. The samples were subjected to molecular method for Leishmania species identification. Results: The digestion pattern of restriction enzyme revealed that 37 (84.1%) CL patients were infected with L. major and 7 (15.9%) were infected with L. tropica. They were mostly male than female. More than half of the patients (58%) had multiple lesions, and they were mostly observed on extremities, 34.1% on legs and 29.5% on hands. Le- sions were mostly of wet ulcerative type. Conclusion: Dominancy of L. major provides more evidence that Varamin District probably could be considered as Zoonotic Cutaneous Leishmaniasis (ZCL) areas. More investigation on other epidemiological aspects of disease is needed. [ABSTRACT FROM AUTHOR]

Background: Although many studies had been conducted on various aspects of canine visceral leishmaniasis (CVL) in domestic dogs in the endemic areas of Iran, investigations on CVL in wild canines are rare. Methods: This is a cross-sectional study was conducted from December 2012 to 2013 in northeast of Iran where human VL is endemic. Wild canines were trapped around the areas where human VL cases had been previously identified. Wild canines were collected and examined both clinically and serologically using direct agglutination test (DAT). Microscopically examinations were performed in all the seropositive wild canines for the presence of the amastigote form of Leishmania spp. Some Leishmania sp. which had been isolated from the spleens of wild canines, were examined analyzed by conventional PCR and sequencing techniques using α-tubulin and GAPDH genes. Results: Altogether, 84 wild canines including foxes (Vulpes vulpes, n=21), Jackals (Canis aureus, n=60) and wolves (Canis lupus, n=3) were collected. Four foxes and seven jackals showed anti-Leishmania infantum antibodies with titers of 1:320-1:20480 in DAT. Furthermore, one fox and one jackal were parasitologically (microscopy and culture) positive and L. infantum was confirmed by sequence analysis. Conclusion: The present study showed that sylvatic cycle of L. infantum had been established in the studied endemic areas of VL in northeastern Iran. [ABSTRACT FROM AUTHOR]

Background Resistance to pentavalent antimonial drugs has become a serious problem in the treatment of cutaneous leishmaniasis in some endemic areas. Investigations on molecular markers involved in drug resistance are essential for monitoring of the disease. Leishmania-activated C kinase gene ( LACK1) is involved in multiple central processes such as signal transduction. According to the probable role of the LACK1 gene in antimony resistance, we used real-time reverse transcription-polymerase chain reaction ( PCR) to investigate the expression of this gene in clinical L. tropica strains, which were resistant or sensitive to meglumine antimoniate. Methods We analyzed the expression level of LACK in 18 sensitive and 14 resistant L. tropica isolates collected from patients with anthroponotic cutaneous leishmaniasis. After cDNA synthesis, gene expression analysis was performed by quantitative real-time PCR using SYBR Green. In addition, the full length of the LACK gene from six reference strains was cloned and sequenced then deposited in the NCBI database to confirm our strains. Results Real-time reverse transcription- PCR revealed that the average RNA expression level of LACK in isolates from unresponsive and responsive patients were 0.479 and 4.583, respectively, and expression of LACK was significantly downregulated (9.56-fold) in resistant isolates compared to sensitive ones. Conclusion Results of the present study suggest the probable role of the LACK gene in antimony resistance. Moreover, it can be considered as a potential marker for monitoring antimony resistance in clinical isolates. However, further studies are required to exploit the biological functions of it in antimony resistance. [ABSTRACT FROM AUTHOR]

Introduction: Cutaneous leishmaniasis (CL) is among the top 10 infectious disease priorities in the world, and the leading cause of morbidity in Iran. The present study was conducted to assess the risk of CL, and to determine some epidemiological features of the disease in endemic areas of Qom Province in Central Iran during 2009 to 2013. Methods: Data regarding human cases of the disease were obtained from the Qom Province Health Center, prepared and stored in a spatial database created in ArcGIS10.3. A total of 9 out of 212 Leishmania spp. positive slides taken in 2013 from patients residing in Qom city were examined using molecular methods and the species of Leishmania was identified by PCR-RFLP. Those 9 patients had no history of travel outside the city. Spatial analysis and clustering methods were applied to find major hot spots and susceptible areas for the establishment of novel foci of the disease. Transmission patterns were examined for spatial autocorrelation using the Moran's I statistical application, and for the clustering of high or low values using the Getis-Ord Gi* statistics. Results: During the period of study, a total of 1767 CL cases were passively reported in the area, out of which were 65% males and 35% females. The highest and lowest numbers of cases were reported in 2010 and 2013, respectively. Importantly, 979 cases were reported from urban areas, while the remainder came from rural areas. Leishmania major was detected as the causative agent of CL in the city of Qom. Remarkably, most patients recorded in Qom city were associated with a history of travel to the endemic areas of CL within the province, or to other endemic areas of the disease in Iran. Spatial distribution of CL cases revealed northeastern and southwestern quarters of the city were the major hot spots of the disease (P<0.05). Hot spot and CL transmission risk analysis across the province indicated that more than 40 villages were located in high and very high risk areas of CL transmission. Conclusions: Due to the existence of hot spots (P<0.05) of CL in successive years in some quarters of Qom city, along with detection of L. major from the patients without a history of travel, there may be potential of local transmission of the disease within the city. Therefore, it is necessary to conduct a comprehensive study concerning the hot spots of CL in Qom city for curtailing the incidence of the disease in the city. The methodology and the results of this study is essential in serving as a yardstick for subsequent similar studies that will be carried out in other endemic areas of CL in Iran and providing an adequate tool for the establishment of a national database of cutaneous leishmaniasis. [ABSTRACT FROM AUTHOR]

We describe a case of visceral leishmaniasis (VL) due to Leishmania tropica in a 50- year-old Iranian man lived in a VL-endemic area in southwest of Iran. The patient presented with a 3-month history of fever and splenomegaly. Clinical signs and serological findings were suggestive of VL. Spleen biopsy was taken from the patient and intracellular forms of Leishmania amastigotes was seen in Giemsa stained smears. The patient was treated with pentavalent antimonial compound with complete resolution of his systemic signs and symptoms. DNA was extracted from the microscopic slides of the spleen biopsy and the nagt (N-Acetylglucosamine-1- Phosphate Transferase) gene of Leishmania was PCR-amplified. Sequence analysis of the PCR product demonstrated that the case has 99% identity with those of available sequences of L. tropica. Intra-species variation within isolate was 0-0.1%; whereas, inter-species differences of the isolate with those of L. major and L. infantum was significantly higher. [ABSTRACT FROM AUTHOR]