Publications by authors named "Hatam GholamReza"

46 Publications

Designing of a Novel Fusion Protein Vaccine Candidate Against Human Visceral Leishmaniasis (VL) Using Immunoinformatics and Structural Approaches.

Int J Pept Res Ther 2021 Apr 24:1-14. Epub 2021 Apr 24.

Basic Sciences in Infectious Diseases Research Center, Shiraz University of Medical Sciences, Shiraz, Iran.

Leishmaniasis is caused by an obligate intracellular protozoan parasite. The clinical forms of leishmaniasis differ from cutaneous leishmaniasis, mucocutaneous leishmaniasis and visceral leishmaniasis (VL) which depend on the parasite species and the host's immune responses. There are significant challenges to the available anti-leishmanial drug therapy, particularly in severe forms of disease, and the rise of drug resistance has made it more difficult. Currently, no licensed vaccines have been introduced to the market for the control and elimination of VL. A potential target for use in candidate vaccines against leishmaniasis has been shown to be Kinetoplastid membrane protein-11 (KMP-11) antigen. In this study, we chose KMP-11 antigen as target antigen in our vaccine construct. In addition, B-type flagellin (fliC) was used as an adjuvant for enhancing vaccine immunogenicity. The GSGSGSGSGSG linker was applied to link the KMP-11 antigen and fliC (KMP-11-fliC) to construct our fusion protein. Bioinformatics approaches such as; 3D homology modeling, CTL, B-cell, MHC class I and II epitopes prediction, allergenicity, antigenicity evaluations, molecular docking, fast simulations of flexibility of docked complex and in silico cloning were employed to analysis and evaluation of various properties of the designed fusion construct. Computational results showed that our engineered structure has the potential for proper stimulation of cellular and humoral immune responses against VL. Consequently, it could be proposed as a candidate vaccine against VL according to these data and after verifying the efficacy of the candidate vaccine through in vivo and in vitro immunological tests.
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http://dx.doi.org/10.1007/s10989-021-10218-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8067785PMC
April 2021

Genetic immunization against toxoplasmosis: A review article.

Microb Pathog 2021 Apr 27;155:104888. Epub 2021 Apr 27.

Department of Parasitology and Mycology, Shiraz University of Medical Sciences, Shiraz, Iran; Basic Sciences in Infectious Diseases Research Center, Shiraz University of Medical Sciences, Shiraz, Iran. Electronic address:

Toxoplasma gondii is a protozoan coccidian parasite belonging to Phylum Apicomplexa and is the causative agent of toxoplasmosis as a zoonotic disease around the world. It is one of the most important protozoa which is transmitted via various routes and infects several warm-blooded animals. The seroprevalence of T. gondii infection is high worldwide and leads to clinical, psychological, and economic problems. At present, available drug therapy for toxoplasmosis has severe side effects, so the development of new anti-toxoplasma drugs or effective vaccines is mandatory. Therefore, different measures have been taken for the development of anti-toxoplasmosis vaccines, and various studies have shown that DNA vaccines could be one of the most successful approaches against the intracellular parasite, T. gondii. Many of these studies have evaluated the efficacy of immunogenicity and different aspects of the DNA vaccines for toxoplasmosis including single genes or multi-gene plasmids with or without adjuvants. Most of the literature confirms that DNA vaccines containing different antigens of the toxoplasma parasite can induce suitable immune response and protection in acute or chronic toxoplasmosis. Therefore, in this review article, we aimed to discuss the current status of DNA vaccines as a new immunization method against toxoplasmosis.
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http://dx.doi.org/10.1016/j.micpath.2021.104888DOI Listing
April 2021

Immunotherapy in treatment of leishmaniasis.

Immunol Lett 2021 May 23;233:80-86. Epub 2021 Mar 23.

Department of Parasitology, Shiraz University of Medical Sciences, Shiraz, Iran. Electronic address:

Leishmaniasis caused by various species of protozoan transmitted by sand fly vectors occurs as a spectrum of clinical features including cutaneous, mucocutaneous and visceral forms. It is a geographically distributed parasitic disease and a major public health problem in the world. The clinical syndromes are highly variable depending on the parasite species, host genetics, vectors and environment. To date, there is no effective vaccine and traditional treatments are toxic, expensive with long administration duration and many adverse side effects and/or drug resistance. Instead of treatments based on chemotherapy, certain strategies aim to recover leishmaniasis and reduce the parasitic burden. Immunotherapy has focused on the induction of effective immune response to rapidly control the disease. Recent studies have indicated that a single dose of a suitable therapeutic vaccine induces a quick and lasting recovery in patients. Immunotherapy reduces the toxicity of drug and the emergence of resistance dramatically. It could be an effective addition to chemotherapy with a safe and potent drug compared with monotherapy, resulting in a prophylactic and therapeutic cure of leishmaniasis. This review has focused on treatment of leishmaniasis with particular emphasis on immunotherapy as an alternative to conventional drug treatment.
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http://dx.doi.org/10.1016/j.imlet.2021.03.011DOI Listing
May 2021

FML-ELISA a novel diagnostic method for detection of feline leishmaniasis in two endemic areas of Iran.

J Parasit Dis 2021 Mar 10;45(1):279-284. Epub 2020 Nov 10.

Basic Sciences in Infectious Diseases Research Center, Shiraz University of Medical Sciences, Shiraz, Iran.

Although canids are regarded as major reservoir hosts for , feline leishmaniasis are reported sporadically from different endemic foci of Mediterranean visceral leishmaniasis (VL). Despite the risk of parasite transmission between human and other animals, most of the studies are limited to dogs and few studies are focused to investigate sp. among other mammals. This project was aimed to detect antibodies of cats in two VL endemic regions of Iran by Fucose Mannose Ligand (FML) and soluble antigen (SLA) ELISA. Forty nine stray cats of different age and sex, from Fars and Ardabil provinces (two VL endemic loci of Iran) were sampled, then tested for by FML and SLA-ELISA. Sixteen percent (8/49) of cat sera were reported positive by FML-ELISA. SLA-ELISA showed 18.3% (9/48) positive cases in cats. Sensitivity of FML-ELISA was calculated 57% and SLA ELISA 25%. Specificity of FML and SLA ELISA were assessed 78% and 68% respectively. Kappa coefficient of agreement between FML and SLA-ELISA was detected on 0.45. As feline leishmaniasis could be a potential risk in endemic areas, FML-ELISA could be considered as an appropriate examination to detect leishmaniasis in cats.
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http://dx.doi.org/10.1007/s12639-020-01316-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7921240PMC
March 2021

Comparative molecular prevalence and subtypes distribution of Blastocystis sp. a potentially zoonotic infection isolated from symptomatic and asymptomatic patients in Iran: A systematic review and meta-analysis.

Acta Parasitol 2021 Mar 9. Epub 2021 Mar 9.

Basic Sciences in Infectious Diseases Research Center, Shiraz University of Medical Sciences, Shiraz, Iran.

Purpose: The present systematic review and meta-analysis was aimed to assess the weighted molecular prevalence of Blastocystis sp. in humans along with the comparative molecular prevalence and subtypes distribution of Blastocystis isolated from symptomatic and asymptomatic patients in Iran.

Methods: International electronic databases including Medline/PubMed, ProQuest, Scopus, Embase, and Google Scholar were explored until 4th October 2020. Heterogeneity index was evaluated among studies using Cochran's Q test and I index. Finally, 23 eligible studies were qualified to be included in this review.

Results: The pooled molecular prevalence of Blastocystis sp. in Iran was reported 15.2% (95% CI 11.5-19.7). In addition, the molecular prevalence based on PCR-sequencing and STS primers was reported 12.5% (95% CI 8.6-17.7) and 19.8% (95% CI 13.1-28.8), respectively. Interestingly, there was a considerably higher prevalence among asymptomatic patients [25.1% (95% CI 20.8-30.0)] in comparison to symptomatic ones [21.0% (95% CI 15.9-27.2)]. In addition, the frequency of Blastocystis ST1, ST2, and ST3 from positive samples in symptomatic patients was 19.7%, 35.1%, and 47.4%, respectively. In addition, the prevalence of Blastocystis ST1, ST2, and ST3 from positive samples in asymptomatic patients was 27.1%, 26.8%, and 37.8%, respectively. The results obtained in Iran showed that Blastocystis is more common in asymptomatic patients compared to patients having clinical symptoms. Of note, ST3, as the most common subtype causing clinical symptoms, was the most prevalent reported subtype among both symptomatic and asymptomatic patients in the country.

Conclusions: Hence, the pathogenicity of the Blastocystis parasite is not subtype-specific and appears to be related to a variety of risk factors. Still the Blastocystis epidemiology is open to question and more large-scale studies should be performed on this aspect.
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http://dx.doi.org/10.1007/s11686-021-00360-0DOI Listing
March 2021

The Efficacy of Hydroalcoholic Extracts of Prosopis farcta Against

Turkiye Parazitol Derg 2021 03;45(1):1-4

Birjand University of Medical Sciences, Department of Public Health, Birjand, İran

Objective: Cutaneous leishmaniasis caused by () is an endemic disease in Iran. The current reference drugs, including Glucantime, possess high toxicity in addition to some side-effects. Therefore, there is a growing interest in exploring biomedical plants. The goal of the present study was to evaluate the anti-leishmanial activity and cytotoxicity of hydroalcoholic extracts from () over promastigote and amastigote forms.

Methods: This study was performed at the Iran Birjand University of Medical Sciences, during the year 2019. In this study, the hydroalcoholic extracts of the stems, leaves (LE) and fruits (FE) of were obtained. The anti-leishmanial activity was assessed against leptomonad promastigotes and intracellular amastigotes of . The cytotoxicity of these extracts was determined in murine macrophages.

Results: The FE and LE of demonstrated a significant leishmanicidal effect against promastigotes with an IC50 of 0.9 mg/mL and 1.1 mg/mL, respectively. The FE showed the most anti-leishmanial activity and presented with the highest index of selectivity (SI=14.6) as an anti-leishmanial product. Infected macrophages treated using the FE showed a reduction in parasite burden by 97.3%.

Conclusion: The results of the present study demonstrated the leishmanicidal activity of on both promastigotes and intracellular amastigotes. There is a need for performing comprehensive studies on relevant animal models and to access the effects of active components of extract on the growth of L. major.
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http://dx.doi.org/10.4274/tpd.galenos.2020.6956DOI Listing
March 2021

Potential therapeutic targets shared between leishmaniasis and cancer.

Parasitology 2021 May 4;148(6):655-671. Epub 2021 Feb 4.

Department of Microbiology and Parasitology, IdiSNA (Navarra Institute for Health Research), c/ Irunlarrea 1, University of Navarra, ISTUN Instituto de Salud Tropical, 31008Pamplona, Spain.

The association of leishmaniasis and malignancies in human and animal models has been highlighted in recent years. The misdiagnosis of coexistence of leishmaniasis and cancer and the use of common drugs in the treatment of such diseases prompt us to further survey the molecular biology of Leishmania parasites and cancer cells. The information regarding common expressed proteins, as possible therapeutic targets, in Leishmania parasites and cancer cells is scarce. Therefore, the current study reviews proteins, and investigates the regulation and functions of several key proteins in Leishmania parasites and cancer cells. The up- and down-regulations of such proteins were mostly related to survival, development, pathogenicity, metabolic pathways and vital signalling in Leishmania parasites and cancer cells. The presence of common expressed proteins in Leishmania parasites and cancer cells reveals valuable information regarding the possible shared mechanisms of pathogenicity and opportunities for therapeutic targeting in leishmaniasis and cancers in the future.
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http://dx.doi.org/10.1017/S0031182021000160DOI Listing
May 2021

Identification of immunoreactive proteins in secretions of Leishmania infantum promastigotes: an immunoproteomic approach.

East Mediterr Health J 2020 Dec 9;26(12):1548-1555. Epub 2020 Dec 9.

Basic Sciences in Infectious Diseases Research Center, Shiraz University of Medical Sciences, Shiraz, Islamic Republic of Iran.

Background: In the Mediterranean region, Leishmania infantum is the main cause of visceral leishmaniasis. Dogs with canine visceral leishmaniasis are an important reservoir of visceral leishmaniasis. Control of canine visceral leishmaniasis could disrupt transmission of visceral leishmaniasis to humans. The secreted antigens of Leishmania promastigotes are potential stimuli of the host immune system. Proteomic techniques facilitate the identification of new protein markers.

Aims: This study aimed to identify immunoreactive proteins in the secretions of L. infantum promastigotes which could be possible targets for the diagnosis and treatment of canine visceral leishmaniasis and the development of vaccines against the disease.

Methods: Secretions of L. infantum promastigotes were obtained from the cultivation of 6 × 10 promastigotes in serum- free RPMI-1640 medium during a period of 72 h. After deionization and lyophilization, two-dimensional gel electrophoresis was used for protein separation followed by Western blotting. Thirteen common and repeatable immunoreactive spots were analysed by mass spectrometry.

Results: Nine proteins were identified by spectrometry. Most of these proteins were involved in metabolism pathways, survival and pathogenicity of Leishmania parasites. Phospholipase C, immune inhibitor A, chitin-binding protein and a single peptide match to chain A crystal structure of selenomethionine were observed in the secretions of L. infantum promastigotes.

Conclusions: The proteins identified in metabolism pathways, survival and pathogenicity of Leishmania parasites are possible targets that could be used for the diagnosis and treatment of canine visceral leishmaniasis and the development of vaccines against the disease in the future.
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http://dx.doi.org/10.26719/emhj.20.114DOI Listing
December 2020

Molecular and serological study on congenital toxoplasmosis in newborn of Shiraz, Southern Iran.

Environ Sci Pollut Res Int 2021 Apr 28;28(13):16122-16128. Epub 2020 Nov 28.

Basic Sciences in Infectious Diseases Research Center, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran.

Toxoplasmosis is a zoonotic disease caused by an obligatory intracellular parasite, Toxoplasma gondii. The congenital form of the disease is a significant health problem around the world. This study aimed to determine the incidence of congenital toxoplasmosis in the newborn of Shiraz, Southern Iran, between 2013 and 2018. A total of 2498 blood samples of neonates were randomly collected by a standard heel puncture technique and dried on Guthrie cards from Shiraz newborn screening center. We provided questionnaire forms for mothers according to their demographic characteristics, such as age, place of residence, history of having cats as pet, and literacy level. ELISA immunocapture and PCR assays were applied to detect anti-Toxoplasma IgM and the parasite DNA in dried blood spot samples. The anti-Toxoplasma IgM antibody was detected in two out of the 2498 infants. Moreover, borderline titers were observed in 3 samples, which were considered suspicious, so these were retested after 18 months to detect IgG against Toxoplasma. Positive IgG titer was observed in two infants who had a positive IgM level. The genome of Toxoplasma was detected in one sample out of 2498. No significant differences were seen between the epidemiological factors with congenital infection. The incidence of congenital toxoplasmosis is estimated at 0.08% of studied samples. It seems that this incidence could present to the health ministry as a logical research achievement for the national screening program of newborns in Iran.
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http://dx.doi.org/10.1007/s11356-020-11707-xDOI Listing
April 2021

A proteomic glimpse into the effect of antimalarial drugs on Plasmodium falciparum proteome towards highlighting possible therapeutic targets.

Pathog Dis 2021 01;79(1)

Basic Sciences in Infectious Diseases Research Center, Shiraz University of Medical Sciences, Shiraz, Iran.

There is no effective vaccine against malaria; therefore, chemotherapy is to date the only choice to fight against this infectious disease. However, there is growing evidences of drug-resistance mechanisms in malaria treatments. Therefore, the identification of new drug targets is an urgent need for the clinical management of the disease. Proteomic approaches offer the chance of determining the effects of antimalarial drugs on the proteome of Plasmodium parasites. Accordingly, we reviewed the effects of antimalarial drugs on the Plasmodium falciparum proteome pointing out the relevance of several proteins as possible drug targets in malaria treatment. In addition, some of the P. falciparum stage-specific altered proteins and parasite-host interactions might play important roles in pathogenicity, survival, invasion and metabolic pathways and thus serve as potential sources of drug targets. In this review, we have identified several proteins, including thioredoxin reductase, helicases, peptidyl-prolyl cis-trans isomerase, endoplasmic reticulum-resident calcium-binding protein, choline/ethanolamine phosphotransferase, purine nucleoside phosphorylase, apical membrane antigen 1, glutamate dehydrogenase, hypoxanthine guanine phosphoribosyl transferase, heat shock protein 70x, knob-associated histidine-rich protein and erythrocyte membrane protein 1, as promising antimalarial drugs targets. Overall, proteomic approaches are able to partially facilitate finding possible drug targets. However, the integration of other 'omics' and specific pharmaceutical techniques with proteomics may increase the therapeutic properties of the critical proteins identified in the P. falciparum proteome.
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http://dx.doi.org/10.1093/femspd/ftaa071DOI Listing
January 2021

Comparison of camel, dog and the laboratory animals' sera with the fetal calf serum (FCS) for cultivation of .

J Parasit Dis 2020 Jun 8;44(2):299-304. Epub 2020 Feb 8.

Basic Sciences in Infectious Diseases Research Center, Shiraz University of Medical Sciences, Shiraz, Iran.

The best-known serum for spp. cultivation is the fetal calf serum (FCS), which is very expensive with ethical concerns. This study was conducted to compare various laboratory (BALB/c mice, rat, rabbit, hamster and guinea pig) and non-laboratory (dog and camel) animals' sera as a substitute for FCS in culture. , /// strain, was cultivated in RPMI-1640 medium enriched with different percentages of mentioned animal's sera. Parasite growth was checked constantly. The rate of growth and survival of parasites were compared with a control medium enriched with FCS. As well, biochemical (albumin, globulin AST, ALT, ALP, Bil, BUN, Crea, Ca, P, Na, K, Fe, TIBC, Mg, zinc, Chol, HDL, LDL, TG, BS, uric acid, LDH, CPK) analysis of all sera was performed and compared with FCS. The most promastigote growth rate is considered in 10% BALB/c, guinea pig and hamster sera on the 6th day of cultivation. Also, on the 8th day, parasites showed viability in all animal sera. The promastigote growth in culture media enriched with the camel and the dog sera in comparison with laboratory animals was considered very low. Differences between 10% FCS and 10% cocktail serum were not significant ( > 0.05) but with other sera were significant ( < 0.05). Also, differences between BALB/c with hamster and guinea pig sera were not significant, respectively ( = 0.07 and  = 0.09). According to the biochemical analysis of all sera, the higher content of iron was detected in the hamster, guinea pig, BALB/c and fetal calf sera. The magnesium and zinc content of guinea pig and BALB/c serum was found to be more than the others and comparable with FCS. The promastigote growth decreased by camel, dog and rat sera orderly. In this study, a rapid increase in parasite growth in media supplemented with hamster, BALB/c and guinea pig sera was considered. It could be suggested to use these sera as a suitable alternative for FCS in molecular biology researches.
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http://dx.doi.org/10.1007/s12639-020-01203-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7244685PMC
June 2020

A New Perspective on the Status of the Intestinal Parasitic Infections in the Rural Areas of Fars Province South of Iran.

Iran J Public Health 2019 Aug;48(8):1518-1522

Basic Sciences in Infectious Diseases Research Center, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran.

Background: Parasitoses are among the most important problems of most countries especially developing countries. We aimed to detect the situation of intestinal parasitic infections in the Farashband district in Fars Province South of Iran and identify influential factors in the escalation of parasitic diseases and to reduce them.

Methods: Overall, 1009 participants from the age of 6 months to 90 years were selected from 3 cities and 15 villages of Farashband district, Fars Province South of Iran from 2015 to 2016. Parasitological methods such as the direct assay method, formalin-ether concentration method, and zinc sulfate flotation were used for diagnosis of worm eggs, cysts, and protozoa trophozoite. Susceptible and protozoan positive samples were stained using the Trichrome staining method. The modified acid-fast staining procedure was conducted for diarrheal samples and the results were used for diagnosis of coccidia.

Results: Overall, 313 subjects were infected with at least one intestinal parasite (pathogenic and nonpathogenic). Helminthes infection and protozoan infection were observed in 9 (0.9%) and 304 (30.13%) participants, respectively. Fecal samples of 34 patients with diarrheal feces were used to prepare smears for further examinations using the Ziehl-Neelsen staining method. Examinations showed no infection with coccidia.

Conclusion: Helminthes infection has decreased drastically but protozoan infection is still considered a health issue in this region. It is possible to reduce parasitic infections through proper measures such as increasing public awareness and education the public, especially children on health problems with education courses.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7145925PMC
August 2019

Leishmanial selenoproteins and the host immune system: towards new therapeutic strategies?

Trans R Soc Trop Med Hyg 2020 07;114(7):541-544

Basic Sciences in Infectious Diseases Research Center, Shiraz University of Medical Sciences, Shiraz, 7134845794, Iran.

Optimum levels of selenoproteins are essential for starting and managing the host immune responses against pathogens. According to the expression of selenoproteins in Leishmania parasites, and since high levels of selenoproteins lead to adverse effects on immune cells and their functions, Leishmania parasites might then express selenoproteins such as selenomethionine in their structure and/or secretions able to challenge the host immune system. Finally, this adaptation may lead to evasion of the parasite from the host immune system. The expression of selenoproteins in Leishmania parasites might then induce the development of infection. We therefore suggest these molecules as new therapeutic candidates for the treatment of leishmaniasis.
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http://dx.doi.org/10.1093/trstmh/traa013DOI Listing
July 2020

Molecular identification and subtypes distribution of Blastocystis sp. isolated from children and adolescent with cancer in Iran: evaluation of possible risk factors and clinical features.

Acta Parasitol 2020 Jun 2;65(2):462-473. Epub 2020 Mar 2.

Department of Medical Parasitology and Mycology, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran.

Purpose: This study aimed to determine the molecular characterization and subtype distribution of Blastocystis sp. isolated from cancer children and adolescents in Shiraz, Fars province, southwestern Iran.

Methods: Overall, 200 fecal samples obtained from cancer children and adolescents under 18 years old (107 males and 93 females) and checked by microscopy, culture, and molecular methods (PCR). Possible etiological factors and clinical characteristics of Blastocystis infection were also evaluated and compared between Blastocystis infected and non-infected patients.

Results: Thirteen of 200 (6.5%) stool samples were positive for Blastocystis by microscopy. While 21 of 200 (10.5%) were positive by culture, and 24 of 200 (12%) were positive by PCR. Out of 24 positive samples tested by PCR and sequencing, ST3 was reported as the most common subtype (nine samples, 37.5%), followed by ST2 (eight samples, 33.3%), ST1 (five samples, 20.9%), and ST7 (two samples, 8.3%). The prevalence of Blastocystis infection in males was significantly higher than females (p = 0.024). Also, Blastocystis was more prevalent in patients who had received at least eight chemotherapy cycles than fewer (p = 0.002). However, no associations were found between Blastocystis-positive rate and age, residence, type of cancers, or contact with animals. Also, there was no significant difference between frequency of Blastocystis subtypes in symptomatic and asymptomatic cancer patients.

Conclusions: Various controlled epidemiologic and topographic studies need to confirm or reject these possible associations with Blastocystis infection. The data from this study are an invaluable addition to the growing body of research studies on Blastocystis infection in cancer patients.
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http://dx.doi.org/10.2478/s11686-020-00186-2DOI Listing
June 2020

Chitin binding protein as a possible RNA binding protein in Leishmania parasites.

Pathog Dis 2020 02;78(1)

Basic Sciences in Infectious Diseases Research Center, Shiraz University of Medical Sciences, Shiraz, Iran.

Leishmaniasis includes a broad spectrum of pathological outcomes in humans caused by protozoan parasites from the genus Leishmania. In recent years, proteomic techniques have introduced novel proteins with critical functions in Leishmania parasites. Based on our report of a Chitin binding protein (CBP) in our previous immunoproteomic study, this article suggests that CBP might be an RNA binding protein (RBP) in Leishmania parasites. RBPs, as key regulatory factors, have a role in post-transcriptional gene regulation. The presence of RBPs in Leishmania parasites has not been considered so far; however, this study aims to open a new venue regarding RBPs in Leishmania parasites. Confirming CBP as an RBP in Leishmania parasites, exploring other RBPs and their functions might lead to interesting issues in leishmaniasis. In fact, due to the regulatory role of RBPs in different diseases including cancers and their further classification as therapeutic targets, the emerging evaluation of CBP and RBPs from Leishmania parasites may allow the discovery of novel and effective drugs against leishmaniasis.
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http://dx.doi.org/10.1093/femspd/ftaa007DOI Listing
February 2020

Lipophilic tracer Dil and fluorescence labeling of acridine orange used for tracing in the fibroblast cells.

Heliyon 2019 Dec 18;5(12):e03073. Epub 2019 Dec 18.

Basic Sciences in Infectious Diseases Research Center, Shiraz University of Medical Sciences, Shiraz, Iran.

Background: This study aims to evaluate the use of fluorescent dye Dil and super vital dye acridine orange (AO) tracking of labeled in the fibroblast cells.

Methods: Dil crystal and AO were used to stain in a co-culture of the fibroblasts with the parasite. AO staining solution was added to 1 × 10 parasites. After 10 min, the stained parasites were centrifuged and washed seven times with phosphate buffered saline (PBS). The stained promastigote was incubated with fibroblasts for 6-8 h. The presence of stained parasites with AO in the fibroblast was assessed using a fluorescence microscope. 1 × 10/mL promastigote of was gently suspended and mixed by Dil staining solution with an ultimate concentration of 0.002 μg/mL and it was kept for 20 min at the room temperature. Subsequently, after washing it in PBS for seven times, it was centrifuged at 3000 rpm for 10 min. The supernatant was removed and the precipitate containing stained promastigote was suspended in fresh DMEM F12 with fibroblasts at 37 °C for 6 h. The presence of stained parasites with Dil in fibroblast was assessed using a fluorescence microscope. Fibroblast characterization was undertaken by a real-time polymerase chain reaction (PCR).

Results: Acridine orange staining assisted in detection of the live parasite in the fibroblast cells. Free promastigote looked green before entering into the fibroblasts after 12 h culture. The parasite entered the cytoplasm of fibroblasts at the beginning of the exposure and gradually entered the nucleus of the fibroblast. The fibroblast nucleus was entirely stained green by AO. The appeared green under the fluorescent microscope. Dil staining revealed that the internalized parasites with red/orange color were localized within the cytoplasm after 6-hours and the nucleus of the fibroblasts after 72-hours following culture. Human fibroblasts were positive at the expression of CD10, CD26, matrix metalloproteinase-1 (MMP-1) and matrix metalloproteinase-3 (MMP-3) and negative for CD106 and integrin alpha 11.

Conclusion: The fluorescent dye Dil staining is a safe, easy to use, inexpensive and fast method for labeling of the parasite in the fibroblast cells. Acridine orange staining could be useful for tracing the parasites in the fibroblasts too. In this study, both Dil and AO were compared and considered as suitable vital dyes for identifying labeled in the fibroblast , but Dil was superior to AO with its feature does not transfer from the labeled to unlabeled cells.
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http://dx.doi.org/10.1016/j.heliyon.2019.e03073DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6928280PMC
December 2019

Molecular Identification of Species Caused Cutaneous Leishmaniasis in Southern Zone of Iran.

J Arthropod Borne Dis 2019 Jun 24;13(2):198-205. Epub 2019 Jun 24.

Diagnostic Lab of Leishmaniasis, Valfajr Health Center, Shiraz University of Medical Sciences, Shiraz, Iran.

Background: and are two main species causing cutaneous leishmaniasis (CL) in Iran. Recently, spp. has also been reported in the wound of patients with CL. In this study, we determined the species causing CL in the southern of Iran and the role of spp. in creating skin ulcers.

Methods: In this cross-sectional study from Apr to Sep 2016, 66 patients with CL referred to Diagnostic Lab of Leishmaniasis, Valfajr Health Center, Shiraz, Iran, were selected. After DNA extraction from the Giemsa stained smears, all samples were amplified in two separate steps using specific primers, firstly, to differentiate species and then to identify spp.

Results: Two species and were responsible for 60 and 6 cases, respectively. Moreover, in two patients, mixed infection with was confirmed. In mix infection cases, the morphology of the cutaneous ulcers was not different from the wounds of other patients.

Conclusion: is responsible for the most common CL in southern Iran. In addition, in two patients with and , mix infection with was confirmed. The potential role of as the main factor for CL and the probability of this parasite to have synergistic effects on , as a hypothesis, requires more comprehensive researches on the ambiguity of this protozoon.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6885138PMC
June 2019

CRISPR System: A High-throughput Toolbox for Research and Treatment of Parkinson's Disease.

Cell Mol Neurobiol 2020 May 26;40(4):477-493. Epub 2019 Nov 26.

Department of Tissue Engineering and Applied Cell Sciences, School of Advanced Medical Sciences and Technologies, Shiraz University of Medical Sciences, Shiraz, Iran.

In recent years, the innovation of gene-editing tools such as the CRISPR/Cas9 system improves the translational gap of treatments mediated by gene therapy. The privileges of CRISPR/Cas9 such as working in living cells and organs candidate this technology for using in research and treatment of the central nervous system (CNS) disorders. Parkinson's disease (PD) is a common, debilitating, neurodegenerative disorder which occurs due to loss of dopaminergic neurons and is associated with progressive motor dysfunction. Knowledge about the pathophysiological basis of PD has altered the classification system of PD, which manifests in familial and sporadic forms. The first genetic linkage studies in PD demonstrated the involvement of Synuclein alpha (SNCA) mutations and SNCA genomic duplications in the pathogenesis of PD familial forms. Subsequent studies have also insinuated mutations in leucine repeat kinase-2 (LRRK2), Parkin, PTEN-induced putative kinase 1 (PINK1), as well as DJ-1 causing familial forms of PD. This review will attempt to discuss the structure, function, and development in genome editing mediated by CRISP/Cas9 system. Further, it describes the genes involved in the pathogenesis of PD and the pertinent alterations to them. We will pursue this line by delineating the PD linkage studies in which CRISPR system was employed. Finally, we will discuss the pros and cons of CRISPR employment vis-à-vis the process of genome editing in PD patients' iPSCs.
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http://dx.doi.org/10.1007/s10571-019-00761-wDOI Listing
May 2020

Amino acid mutation in dihydrofolate reductase and dihydropteroate synthetase genes in Hormozgan Province, southern Iran.

J Vector Borne Dis 2019 Apr-Jun;56(2):170-173

Basic Sciences in Infectious Diseases Research Center, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran.

Molecular analysis of antifolate resistance-associated genes-dihydrofolate reductase (dhfr) and dihydropteroate synthetase (dhps) of Plasmodium vivax is important in predicting the emergence of drug resistance to sulphadoxine-pyrimethamine (SP). The present study aimed to determine the polymorphism of dhfr and dhps genes in P. vivax field isolates. Samples from 80 microscopically diagnosed vivax malaria cases were collected from endemic areas of malaria in Hormozgan Province of Iran, from June 2010 to November 2015. The two sets of codons at position 33, 57, 58, 117, 173 of dhfr and 382, 383, and 553 of dhps genes were analysed by direct sequencing of PCR products. The majority of the isolates (70%) harboured a wild-type allele for P. vivax dhfr (Pvdhfr) and P. vivax dhps (Pvdhps). Mutations were detected in three codons of Pvdhfr (P33L, S58R and S117N) and single codon in Pvdhps (A383G). Novel mutations that have not been identified previously at codon 459 (D459A) of Pvdhps were also observed. The high prevalence of point mutation as well as the rising triple mutation of Pvdhfr and Pvdhps genotypes necessitate change in programmes and guidelines to eliminate P. vivax in future.
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http://dx.doi.org/10.4103/0972-9062.263716DOI Listing
December 2019

Cytokine profile and nitric oxide levels in macrophages exposed to Leishmania infantum FML.

Exp Parasitol 2019 Aug 22;203:1-7. Epub 2019 May 22.

Cellular and Molecular Research Center, Yasuj University of Medical Sciences, Yasuj, Iran; Student Research Committee, Yasuj University of Medical Sciences, Yasuj, Iran.

Fucose-mannose ligand (FML) is a soluble antigen purified from Leishmania donovani complex and used for diagnosis, prognosis, and vaccine development against visceral leishmaniasis (VL). We aimed to explore the effects of FML on the production of cytokines, chemokines and nitric oxide (NO) by macrophages in vitro. Peritoneal macrophages from BALB/c mice were treated with various concentrations of FML purified from Leishmania infantum in the absence or presence of LPS Peritoneal macrophages. After 48hr, cell culture supernatants were recovered and the levels of TNF-α, IL-10, IL-12p70 and IP-10 measured by Sandwich ELISA and NO concentration by Griess reaction. We found that FML significantly increase NO, IL-12p70 and IP-10 production in both LPS-treated and untreated macrophages and increase IL-10 levels only in LPS-treated macrophages. However, FML could not alert TNF-α levels in both LPS-treated and untreated macrophages. Further analysis revealed that FML can also increase IL-12p70/IL-10 ratio in LPS-treated macrophages. We concluded that FML can polarize macrophages to an appropriate phenotype similar to M1 phenotype against Leishmania donovani complex, although IL10 and TNF results are controversial.
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http://dx.doi.org/10.1016/j.exppara.2019.05.004DOI Listing
August 2019

An immunoproteomic approach to identifying immunoreactive proteins in amastigotes using sera of dogs infected with canine visceral leishmaniasis.

Pathog Glob Health 2019 05 17;113(3):124-132. Epub 2019 May 17.

c Basic Sciences in Infectious Diseases Research Center, Shiraz University of Medical Sciences , Shiraz , Iran.

Visceral leishmaniasis (VL), the most severe form of leishmaniasis, is caused by and . The infected dogs with canine visceral leishmaniasis (CVL) are important reservoirs for VL in humans, so the diagnosis, treatment and vaccination of the infected dogs will ultimately decrease the rate of human VL. Proteomics and immunoproteomics techniques have facilitated the introduction of novel drug, vaccine and diagnostic targets. Our immunoproteomic study was conducted to identify new immunoreactive proteins in amastigote form of . The strain of (MCAN/IR/07/Moheb-gh) was obtained from CVL-infected dogs. J774 macrophage cells were infected with the promastigotes. The infected macrophages were ruptured, and pure amastigotes were extracted from the macrophages. After protein extraction, two-dimensional gel electrophoresis was employed for protein separation followed by Western blotting. Western blotting was performed, using symptomatic and asymptomatic sera of the infected dogs with CVL. Thirteen repeatable immunoreactive spots were identified by Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Some, including prohibitin, ornithine aminotransferase, annexin A4, and apolipoprotein A-I, have been critically involved in metabolic pathways, survival, and pathogenicity of parasites. Further investigations are required to confirm our identified immunoreactive proteins as a biomarker for CVL.
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http://dx.doi.org/10.1080/20477724.2019.1616952DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6586132PMC
May 2019

Study of multidrug resistance in prevalent Gram-negative bacteria in burn patients in Iran: A systematic review and meta-analysis.

J Glob Antimicrob Resist 2019 12 4;19:64-72. Epub 2019 May 4.

Burn and Wound Healing Research Center, Surgical Department, Shiraz University of Medical Sciences, Shiraz, Iran.

Objectives: Recently, multidrug-resistant (MDR) Gram-negative bacteria (GNB) have become a serious concern causing infections in hospitalised burn patients. This meta-analysis was conducted to detect the prevalence of infections caused by MDR-GNB in hospitalised burn patients in Iran.

Methods: An electronic search was performed using PubMed, Scopus, Web of Science, EMBASE and Iranian databases. Statistical analysis was performed using STATA13. According to the results of the heterogeneity test, a fixed- or random-effects model was used. Publication bias was detected based on Egger's test. Of 1292 articles identified in the initial search, 107 studies were included in this review.

Results: According to the results, the lowest resistance rate was observed in Acinetobacter baumannii and Pseudomonas aeruginosa to colistin, estimated at 21% [95% confidence interval (CI) 2-49%; I=97.30%] and 27% (95% CI 0-82%; I=99.27%), respectively. Moreover, the highest rate was to cefepime, estimated 98% (95% CI 93-100%; I=88.28%) and 87% (95% CI 81-92%; I=96.27%).

Conclusion: The prevalence of MDR-GNB in Iranian burn patients is very high. Thus, a comprehensive infectious control programme, a reduction in the use of antibiotic prophylaxis, and thorough information regarding antimicrobial resistance patterns is required.
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http://dx.doi.org/10.1016/j.jgar.2019.04.017DOI Listing
December 2019

The Importance of Checking Leishmania Promastigotes Viability in the Proteomics Analysis of Secretions.

Turkiye Parazitol Derg 2018 Dec;42(4):245-248

Basic Sciences in Infectious Diseases Research Center, Shiraz University of Medical Sciences, Shiraz, Iran.

Objective: The aim of the present study was to compare the efficacy of checking the viability of Leishmania promastigote by flow cytometry using propidium iodide (PI) and microscopic method using trypan blue (TB) before proteomics analysis of the secretions.

Methods: The promastigotes (6×109) of Leishmania infantum in the exponential growth phase were transferred to serum-free media. Then, the viability of promastigotes was checked and compared with flow cytometry and microscopic method at 0, 2, 3, 4, 5, and 72 h.

Results: Flow cytometry did not show many dead cells at 0 to 4 h, and the viability was approximately 98%. The percentage of the dead promastigotes increased to 8% at 5 h and 17% at 72 h. Meanwhile, the microscopic method using TB did not show any dead cell after 4 and 72 h, and the viability was 100%.

Conclusion: The present study confirms the importance of flow cytometry using PI in checking the viability of Leishmania promastigotes, especially before the proteomics analysis of the secretions. It also shows that flow cytometry using PI is more sensitive than microscopic method using TB.
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http://dx.doi.org/10.5152/tpd.2018.5834DOI Listing
December 2018

Novel Catechol Derivatives of Arylimidamides as Antileishmanial Agents.

Chem Biodivers 2018 Oct 24;15(10):e1800228. Epub 2018 Sep 24.

Basic Sciences in Infectious Diseases Research Center, Shiraz University of Medical Sciences, Shiraz, Iran.

Two novel bis-arylimidamide derivatives with terminal catechol moieties (9a and 10a) and two parent compounds with terminal phenyl groups (DB613 and DB884) were synthesized as dihydrobromide salts (9b and 10b). The designed compounds were hybrid molecules consisting of a catechol functionality embedded in an arylimidamide moiety. All compounds were examined for in vitro antiparasitic activity upon promastigotes of Leishmania major and L. infantum as well as axenic amastigotes of L. major. It was shown that conversion of terminal phenyl groups into catechol moieties resulted in more than 10-fold improvement in potency, coupled with lower cytotoxicity against fibroblast cells, compared to the corresponding parent compounds. The furan-containing analog 9a exhibited the highest activity with submicromolar IC values, ranging from 0.29 to 0.36 μm, which is comparable in efficacy to the reference drug amphotericin B (IC 0.28 - 0.33 μm). The results justify further study of this class of compounds. It seems that the combination of catechol chelating groups with potent antiparasitic agents could improve the efficacy by presenting novel hybrid compounds.
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http://dx.doi.org/10.1002/cbdv.201800228DOI Listing
October 2018

Achievement amastigotes of and investigation of pathological changes in the tissues of infected golden hamsters.

J Parasit Dis 2018 Jun 5;42(2):187-195. Epub 2018 Mar 5.

3Basic Sciences in Infectious Diseases Research Center, Shiraz University of Medical Sciences, Shiraz, Iran.

is an agent of visceral leishmaniasis (VL). Amastigote form is a more appropriate target for investigations on vaccines, treatment, and diagnosis. This study aimed to achieve the amastigotes of in the golden hamster and J774 macrophages and report the pathological changes that occur in the liver and spleen of the hamsters with VL. 4 male golden hamsters were infected with promastigotes. After 5 months, the hamsters were euthanized and touch and pathology smears were prepared from the livers and spleens. Then, these tissues were homogenized and centrifuged at 100×. Supernatants were collected and centrifuged at 2000× and the pellets were collected. In the next part of our study, J774 macrophages were infected with promastigotes. Then, the infected macrophages were ruptured. Centrifuge stages were done same the previous part. The amastigotes were observed in touch and pathology smears. A load of amastigotes in the livers was more than the spleens in both types of smears. Although the livers' structure had undergone pathological changes, the spleens were unchanged. Also, the macrophage infectivity ratio was up to 95%. Our results present a simple and accessible way of achieving a lot of pure and real amastigotes for different fields in Also, it seems that the pathological changes occurring in the spleen and the liver of animals with VL are different and probably can be attributed to the genetic and immune process of the infected animals.
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http://dx.doi.org/10.1007/s12639-018-0981-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5962490PMC
June 2018

Using proteomics as a powerful tool to develop a vaccine against Mediterranean visceral leishmaniasis.

J Parasit Dis 2018 Jun 19;42(2):162-170. Epub 2018 Mar 19.

3Basic Sciences in Infectious Diseases Research Center, Shiraz University of Medical Sciences, Shiraz, Iran.

Visceral leishmaniasis (VL) is a tropical infectious disease, which is called Mediterranean visceral leishmaniasis (MVL) in the Mediterranean area. In spite of many attempts, no effective commercial vaccine exists for MVL. To find new targets for developing antileishmanial vaccines, knowing parasite antigens that provoke the immune system are on demand. Nowadays, proteomics methods are defined as approaches for analysis of protein profiling of different cells. Within this framework, detection of new antigens is becoming more facilitated. In this review, we aimed to introduce possible targets using proteomics so; they could be used as candidates for developing vaccines against MVL. It can shed new light in the near future on the development of promising vaccines for MVL.
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http://dx.doi.org/10.1007/s12639-018-0986-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5962495PMC
June 2018

Phylogenetic Analysis of Human Genotypes in Fars Province, Southern Iran.

Iran J Parasitol 2017 Oct-Dec;12(4):522-533

Dept. of Medical Microbiology and Parasitology, Faculty of Medicine and Health Sciences, University Putra Malaysia, Serdang, Malaysia.

Background: This study is the first phylogenetic genotype analysis of in Iran. The main objective was to determine genotyping and identify the sub-assemblages of isolates involved in the transmission of giardiasis in Fars Province, south of Iran, in 2012.

Methods: Forty isolates were collected from the patient's fecal samples with gastrointestinal discomfort referred to the health centers and hospitals in Shiraz, Fars Province, south of Iran. Purification of cysts from fecal samples and DNA extraction were performed using monolayer of sucrose density gradient and Phenol-Chloroform-Isoamylalcohol (PCI) respectively. Semi-nested PCR and sequence analysis were then performed using the primers (GDHeF, GDHiF, and GDHiR) which amplified a 432-bp fragment of glutamate dehydrogenase () gene. Phylogenetic analysis was carried out using a neighbor-joining tree composed of the nucleotide sequences of isolates obtained in this study and the known sequences isolates published in GenBank.

Results: sub-assemblage AII was the most prevalent genotype with 80% of the cases and 20% of the cases belong to sub-assemblage BIII and BIV based on the DNA sequence of the . isolates at Fars Province were widely distributed within assemblage A cluster (sub-assemblage AII) and the remaining isolates were dispersed throughout the assemblage B cluster (sub-assemblage BIII and BIV).

Conclusion: PCR Sequencing and phylogenetic analysis was a proper molecular method for genotyping and discriminating of the of sub-assemblages in fecal samples, using the glutamate dehydrogenase gene that suggests a human contamination origin of giardiasis.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5756302PMC
January 2018

Cytokine Profile of Leishmania Infantum Fucose-Mannose Ligand in Vaccinated Dogs in the Northwest of Iran.

Iran J Immunol 2017 Dec;14(4):293-305

Department of Parasitology and Mycology, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran.

Background: Canine visceral leishmaniasis (CVL) caused by Leishmania infantum is endemic in the northwest and south of Iran. An appropriate vaccine can help to prevent and control visceral leishmaniasis in both humans and animals. Few studies have confirmed that the fucose-mannose ligand (FML) antigen of Leishmania donovani produced protective immunity in dogs against CVL.

Objective: To evaluate the immune responses of vaccinated dogs against FML antigen of L. infantum.

Methods: We isolated the FML antigen from native L. infantum and vaccinated the dogs with FML-saponin in an endemic area of VL in Iran to evaluate the immune responses of vaccinated dogs against this antigen.

Results: Our results indicated a significant increase in the expression of IFN-γ, IL-10 and IL-13, but not IL-12A, gene transcripts in PBMCs of FML-saponin vaccinated dogs in comparison with controls. Our findings showed a significant difference in the ratio of IFN-γ/IL-10 mRNA expression in FML-saponin vaccinated dogs in comparison with two control groups. Moreover, a significant level of anti-FML antibodies was detected in serum of vaccinated dogs.

Conclusion: These findings showed that FML-saponin stimulates both Th1 and Th2 immune responses with predominant Th1 and strong humoral immune responses to produce protective immunity against CVL.
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http://dx.doi.org/IJIv14i4A4DOI Listing
December 2017

Efficacy of cryotherapy plus topical Juniperus excelsa M. Bieb cream versus cryotherapy plus placebo in the treatment of Old World cutaneous leishmaniasis: A triple-blind randomized controlled clinical trial.

PLoS Negl Trop Dis 2017 Oct 5;11(10):e0005957. Epub 2017 Oct 5.

Medicinal Plants Processing Research Center, Shiraz University of Medical Sciences, Shiraz, Iran.

Background: Cutaneous leishmaniasis is one of the highly prevalent endemic diseases in the Middle East and North Africa. Many treatment modalities have been recommended for this condition but success rates remain limited. Herbal remedies have also been used for treatment but evidence-based clinical trials with these products are sparse. In-vitro and in-vivo studies have shown the anti-leishmanial and curative effects of extract of fruits and leaves of Juniperus excelsa (J. excelsa). The aim of this study was to determine the efficacy of topical J. excelsa M. Bieb extract as an adjuvant to cryotherapy for the treatment of human CL.

Materials And Methods: This study was designed as a two-arm triple-blind randomized placebo-controlled clinical trial using a parallel design. Seventy-two patients with clinical diagnosis of CL confirmed by leishmania smears were allocated to receive either a topical formulation of leaf of J. excelsa extract (group A) or placebo (group B) for 3 months. Both groups received cryotherapy as baseline standard treatment. Patients were evaluated before and weekly after the intervention was initiated until complete cure.

Results: Overall, 82% of patients in group A, experienced complete cure and 9% of them had partial cure. On the other hand, 34% in group B reported complete cure, while 14% of them had partial cure at the end of treatment protocol with a significant difference between the two groups (P< 0.001). The mean duration to healing of the lesions in patients who received J. excelsa extract was statistically significantly shorter than the placebo group (p = 0.04). No significant side effect was seen in the J. excelsa extract group except for mild to moderate local irritation after a few weeks in a few numbers of patients.

Conclusion: The results of this study showed that topical J. excelsa extract can be used as an adjuvant treatment modality in addition to cryotherapy for accelerating the time to cure in addition to increasing the complete cure rate in CL.

Trial Registration: ClinicalTrials.gov IRCT2015082523753N1.
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http://dx.doi.org/10.1371/journal.pntd.0005957DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5655399PMC
October 2017

Application of nanotechnology in treatment of leishmaniasis: A Review.

Acta Trop 2017 Aug 28;172:86-90. Epub 2017 Apr 28.

Department of Parasitology, Shiraz University of Medical Sciences, Shiraz, Iran.

Leishmaniasis is a neglected tropical disease caused by a protozoan species of the genus Leishmania affecting mostly the developing countries. The disease with current mortality rate of 50,000 deaths per year threatens approximately 350 million people in more than 90 countries all over the world. Cutaneous, mucocutaneous and visceral leishmaniasis are the most frequent forms of the disease. Chemotherapy still relies on the use of pentavalent antimonials, amphotericin B, liposomal amphotericin B and miltefosin. Treatment of leishmaniasis has remained insufficient since the current antileishmanial agents have several limitations including low efficacy, toxicity, adverse side effects, drug-resistance, length of treatment and cost lines. Consequently, there is an immediate requirement to search for new antileishmanial compounds. New drug delivery devices transport antileishmanial drug to the target cell specifically with minimizing the toxic effects to normal cells. This study attempts to present a comprehensive overview of different approaches of nanotechnology in treatment of leishmaniasis.
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http://dx.doi.org/10.1016/j.actatropica.2017.04.029DOI Listing
August 2017