Iranian Journal of Parasitology
Volume:6 Issue: 3, Jul-Sep 2011

Leishmania is a protozoan parasite belonging to the family Trypanosomatidae, which is found among 88 different countries. The parasite lives as an amastigote in vertebrate macro­phages and as a promastigote in the digestive tract of sand fly. It can be cultured in the laboratory us­ing appropriate culture media. Although the sexual cycle of Leishmania has not been observed during the promastigote and amastigote stages, it has been reported by some researchers. Leishma­nia has eukaryotic cell organization. Cell culture is convenient and cost effective, and because posttranslational modifications are common processes in the cultured cells, the cells are used as hosts for preparing eukaryotic recombinant proteins for research. Several transcripts of rDNA in the Leishmania genome are suitable regions for conducting gene transfer. Old World Leishmania spp. has 36 chromosomes, while New World Leishmania spp. has 34 or 35 chromo­somes. The genomic organization and parasitic characteristics have been investigated. Leishmania spp. has a unique genomic organization among eukaryotes; the genes do not have introns, and the chromosomes are smaller with larger numbers of genes confined to a smaller space within the nucleus. Leishmania spp. genes are organized on one or both DNA strands and are transcribed as polycistronic (prokaryotic-like) transcripts from undefined promoters. Regulation of gene expres­sion in the members of Trypanosomatidae differs from that in other eukaryotes. The trans-splic­ing phenomenon is a necessary step for mRNA processing in lower eukaryotes and is observed in Leishmania spp. Another particular feature of RNA editing in Leishmania spp. is that mitochon­drial genes encoding respiratory enzymes are edited and transcribed. This review will discuss the chromosomal and mitochondrial (kinetoplast) genomes of Leishmania spp. as well as the phenome­non of RNA editing in the kinetoplast genome.

The aim of this study was to conduct a sero-epidemiological survey in Meshkinshahr, Arda­bil Province, northwestern Iran to detect the rate of hydatidosis in the city and nearby villages. Literature shows that no such study has been conducted so far. Overall, 670 serum samples were collected from 194 males and 476 females from patients re­ferred to different health centers of the region. All patients filled out a questionnaire and an informed con­sent. Sera were analyzed using indirect-ELISA test. Ten μg /ml antigens (Antigen B derived from hydatid cyst fluid), serum dilutions of 1:500 and conjugate anti-human coombs with 1:10000 dilutions were util­ized to perform the test. Data analysis was conducted using SPSS software ver. 11.5. The seroprevalence of human hydatidosis was 1.79% by ELISA test in the region. This rate for fe­males was 1.68% and males 2.6%, respectively. There was no significant difference as regards all fac­tors studied and the seropositivity. According to job, farmers and ranchmen had the highest rate of infec­tion as 3.17%. The sero-prevalence of infection was 2.6%% in illiterate people which showed the highest rate. As regards residency, urban life showed no significant difference with rural life (1.1% vs. 2.58%). Age group of 69-90 yr old, with 4.62% as prevalence had the highest rate of positivity. Obtained sero-prevalence of hydatidosis shows more or less a resemblance to other cities of Iran, although due to the specific condition of the city we expected more rate of sero-positivity.

To evaluate immunotherapy against canine visceral leishmaniasis, Leishmania ma­jor antigen and heat-killed Mycobacterium vaccae (SRL172) were used as stimulators of immune de­fense mechanisms and the results were compared with standard chemotherapy meglumine antimoni­ate. Nineteen mongrel dogs aging 1-3 years old were used in this experiment. Infection was carried out in 15 out of 19 dogs using L. infantum, isolated from a naturally infected poly-symptomatic dog. All the cases showed positive serologic results by direct agglutination test during 30-60 days following inoculation. In the first group, which was under chemotherapy (GlucantimeR), one of the members showed recurrence of the disease despite rapid effect of the therapeutic protocol. Im­munotherapy using SRL172 caused complete cleaning of the parasite in group 2, but the speed was less than Glucantime. Immunotherapy using L. major antigen combined with M. vaccae in group 3 and combine administration of immunotherapy and chemotherapy in group 4 both were with relapsing of one case in each group. Group 5 and 6 were consisted of positive and negative con­trol dogs, respectively. Immunotherapy seems to be an adjuvant in treatment of canine leishmaniasis but it needs more investigation for final confirmation.

In this study, we used both ITS1 and ITS2 for molecular identification of Fasciola species. The region between 18S and 28S of ribosomal DNA was used in PCR-RFLP method for molecular identification of Fasciola species. Ninety trematodes of Fasciola were collected during abattoir inspection from livers of naturally infected sheep and cattle from Khorasan, East Azerbaijan, and Fars provinces in Iran. After DNA extraction, PCR was performed to amplify region ITS1, 5.8S rDNA, ITS2. To select a suitable restriction enzyme, we sequenced and ana-lyzed the PCR products of F. hepatica and F. gigantica samples from sheep and cattle. Tsp509I fast digest restriction enzyme was selected for RFLP method that caused the separation specifi-cally of Fasciola species. The fragment approximately 1000bp in all of the Fasciola samples was amplified and then digested with the Tsp509I restriction endonuclease. Seventy F. hepatica and 20 F. gigantica were identified of total 90 Fasciola isolates. The new PCR-RFLP assay using Tsp509I restriction enzyme provides a simple, practical, fast, low cost, and reliable method for identification and differentiation of Fasciola isolates.

The protozoan parasites Cryptosporidium spp. and Giardia are known to occur widely in both raw and drinking waters. They are two of the causative agents of waterborne out-breaks of gastroenteritis throughout the world. In the present study, a PCR assay and FA were developed for detection of Cryptosporidium oocysts and Giardia cyst in environmental samples. We have detected Cryptosporidium spp. oocysts and Giardia cysts in seeded and un-seeded environmental water samples by PCR method. Water samples were spiked with oocysts (50, 100,300,500) and filtrated with a 1.2-µm pore size cellulose nitrate and follow by DNA extrac¬tion and purification by QIAamp DNA mini kit. Nested-PCR assay amplified an 850 bp fragment of 18s rRNA gene specific for Cryptosporidium and 435 bp fragment of glutamate dehydrogenase (GDH) target gene for Giardia. Also many river water from north of Iran, be checked by these methods. Cryptosporidium and Giardia DNAs were detected in seeded water sample and Giardia was detected in all 5 water samples from river in north of Iran by nested- PCR and FA. Also in one river water sample, Cryptosporidium was detected. This protocol is effective for detection of these waterborne parasites in treated and untreated water samples. This study can also serve as a platform for further investigations and research water source in Iran.

Fascioliasis is considered as the most important helminthic infection of cattle and sheep. Traditional approaches using morphological and biologic characters cannot cause a cer­tainty in the accurate and precise identification and intra-specific differences of Fasciola spp. In this study, we identified Fasciola species using ITS-1 marker and described genetic variation of each species of the parasite in isolates from Tabriz slaughterhouse in West Azerbaijan Province, north- western Iran. Overall, 100 samples (50 from sheep and 50 from cattle) morphologically detected as Fasciola worms were studied for identification of Fasciola species by PCR-RFLP method and intra-species variation of the parasite using RAPD-PCR technique. A region of approximately 460bp in all samples was successfully amplified. There were no identifiable variations among the size of PCR products. Two and three fragments in samples correspond to F. hepatica and F. gigantica was seen, respectively, through PCR-RFLP method. No difference was seen in digestion pattern according to host (sheep or cattle). Different types of each species of the parasite was observed using RAPD-PCR technique. We could have an estimate of frequency of F. hepatica and F. gigantic and different genotypes of the parasite in isolates from one locality in north- western of Iran. By extension of such studies in future to other animal hosts (buffalo and goat) and including more regions to sam­pling, the reliability of the results and their application for control programs in zoonotic diseases will be increased.

Based on the efficacy of paromomycin ointment and recent ongoing clinical trials of combination of paromomycin and gentamicin, a new physical form of films of the paromomy­cin and gentamicin was prepared and anti-Leishmania activities of the prepared films were as­sessed in vitro and in vivo. Paromomycin 15% and gentamicin 0.5% was incorporated in a film using ethyl cellu­lose and HPMC (Hydroxyl Propyl Methyl Cellulose). In order to assess the drug release and anti-Leishmania activities of the preparation, a clone L. major parasite was established using a set of modified NNN medium without overlay liquid layer. Therapeutic effects of the films were evalu­ated using Balb/c mice model. The mice were inoculated with 2×106 L. major promastigotes (MRHO/IR/75/ER) and then when the lesions developed the mice were randomly divided in 3 groups, 10 mice per group, and treated with either perpetrated films or placebo for 28 days or left untreated. Growth inhibition of cloned promastigotes showed that the films have enough releasing capacity and in vivo system, the films containing paromomycin and gentamicin was able to re­duce the lesion size and induced complete cure in 80% of the mice but relapse was seen in 60% of the cured mice and overall 50% cure rate was seen during 20 weeks period of the study. It seems that the prepared films might be further used in human clinical trials.

In Anzali Lagoon, there are some endemic and exotic fishes. The present study was conducted to compare the parasitic fauna of Blicca bjeorkna, as an endemic fish and Hemicul­ter leucisculus, as an introduced fish to the lagoon. A parasitological investigation was done on 78 specimens of B. bjoerkna and 114 of H. leu­cisculus. The fishes were collected from August 2009 to April 2010 by the electro fishing from Anzali Lagoon. Eleven parasites species were found in 192 fish samples. The prevalence and mean inten­sity of parasites in each host were as follows: Parasites from B. bjorkna were Trichodina perforata (53.85%); Myxobolus musayevi (27.19%, 1±0.79); Dactylogyrus difformis (88.05%, 8±7.24) and D. sphyrna (5.18%, 0.95±0.51), Diplostomum spataceum (98.72%, 9.51±9.01), Post­hodiplostomum cuticula (15.38%, 4.25±2.5), Ripidocotyle sp. (1.28%, 2±0.74); Contracaecum osculatum (17.95%, 1.64±0.79), Philometra rischta (12.8%, 1.4±0.54), and Raphidascaris acus (1.04%, 0.03±0.26). The H. leucisculus were infected with T. perforata (27.19%), D. spataceum (7.89%, 1.33±0.54), Ps. tomentosa (7.02%, 1.62±0.49) and R. acus (0.88%, 3±0.28). B. bjoerkna was presented as a new host for M. musayevi and C. osculatum, while H. leucisculus was intro­duced as a new host for T. perforata and Ps. tomentosa. The prevalence of parasites was significantly more in native fish than that of exotic fish (P<0.05). This reduction in parasitic infection in H. leucisculus may be due to its immune system resistance, well adaptation to the new environment, host-specific limitation for endemic parasites and disability of introduced parasite to complete its life cycle in the new host as well

The study was aimed to show the effect of molecular mechanism of Aqueous Garlic Extract (AGE) on expression of IFNγ and iNOS genes in Leishmania major. Leishmania major promastigotes (MRHO/IR/75/ER) were added to the in-vitro cultured J774 cell line, the cells were incubated for 72 hours. Various concentrations of garlic extract (9.25, 18.5, 37, 74, 148 mg/ml) were added to the infected cells. MTT assay was applied for cellular proliferation. After 72 hours of incubation, supernatants were collected and total RNA was extracted from the infected cells. The express of IFNγ and iNOS genes were studied by RT-PCR method. The colorimetric MTT assay after 3 days of incubation showed cytotoxic effect of garlic extract with an IC50 of 37 mg/ml. In addition, IFNγ and iNOS genes expression by RT-PCR indicated that garlic extract lead to over expression of these genes in J774 cell line infected with L. major. Garlic extract exerts cytotoxic effect on infected J774 cell line. In addition, the hypothesis that garlic can improve cellular immunity with raising the expression of IFNγ and of iNOS genes con­firmed.

Genus Trichostrongylus (Nematoda: Trichostrongylidae) is one of the most impor­tant zoonotic nematodes with wide geographic distribution in the world. The purpose of the present study was to describe morphological and morphometrical characteristics of male Trichostrongylus species, currently prevalent in domestic ruminants of Khuzestan Province, south­west Iran. Gastro-intestinal organs of 1600 sheep, goats, cattle, and buffalos, slaughtered in Khuzestan Province, southwest Iran, were examined for infectivity with Trichostrongylus spe­cies. For examination and measurements of helminthes, Azo-carmine staining was per­formed, followed by camera lucida drawings of morphological characters and measurements of morphometrical criteria with a calibrated microscope. Using valid nematodes systematic keys, almost all the parasites were identified at the level of species. Overall, 114 animals were found infected with at least one species of Trichostrongy­lus. Considering morphological characteristics of male Trichostrongylus, six species were identi­fied including T. colubriformis, T. vitrinus, T. probolorus, T. capricola, T. longispicu­laris and Trichostrongylus sp. Although, compared to the previous decades, currently Trichostrongylus is much less prevalent in the domestic ruminants of the study area, but still different species occur in these animals.

Nosemosis of European honey bee (Apis mellifera) is present in bee colonies world­wide. Until recently, Nosema apis had been regarded as the causative agent of the disease, that causes heavy economic losses in apicultures. Nosema ceranae is an emerging microsporidian para­site of European honeybees, A. mellifera, but its distribution is not well known. Previously, nosemosis in honeybees in Iran was attributed exclusively to N. apis. Six Nosema positive samples (determined from light microscopy of spores) of adult worker bees from one province of Iran (Savadkouh- Mazandaran, northern Iran) were tested to determine Nosema species using previously- developed PCR primers of the 16 S rRNA gene. As it is difficult to distinguish N. ceranae and N. apis morphologically, a PCR assay based on 16 S ribosomal RNA has been used to differentiate N. apis and N. ceranae. Only N. ceranae was found in all samples, indicating that this species present in Iran apiar­ies. This is the first report of N. ceranae in colonies of A. mellifera in Iran. It seems that intensive surveys are needed to determine the distribution and prevalence of N. ceranae in differ­ent regions of Iran.

The aim of this paper was to study the prevalence and intensity of Anisakids lar­vae in the long tail tuna fish captured from Iranian shores of Persian Gulf. Different organs including skin, abdominal cavity, stomach and intestinal contents, stom­ach sub serous tissues, liver, spleen, gonads and 20 grams of muscles of 100 long tail tuna fish (Thannus tonggol) caught from waters of the north parts of Persian Gulf were searched for anisakid nematodes larvae. Twenty grams of around the body cavity muscles were digested in artificial gastric juice. Different organs and digested muscles were examined with naked eyes for the presence of anisakids larvae. The collected larvae were preserved in 70% alcohol containing 5% glycerin, and cleared in lactophenol for identification. Our findings revealed that 89% of fish harbored 3rd stage larvae of Anisakis sp. of which 2% were infected with both Anisakis and Raphidascaris. All inspected organs except that of skin were found to be infected, while stomach sub serous tissues were the most infected organ (80%) followed by abdominal cavity (10%), liver (4%), testicle (3%), stomach contents and spleen (2%) and intestinal contents (1%). Intestine and abdominal cavity were the organs har­bored Raphidascaris sp. Digested muscles were free of parasite. Mean intensity was low for both spe­cies and ranged between 1.5 for Raphidascaris sp. and 3.67 for Anisaki sp. Anisakids larvae especially Anisakis are very prevalent in some fish including tunas of Persian Gulf, and consumption of infected fish if it is not properly cooked may lead to human anisakiasis.

Trichomonas vaginalis is protozoan parasite responsible for trichomoniasis and is more common in high-risk behavior group such as prostitute individuals. Interest in trichomoni­asis is due to increase one's susceptibility to viruses such as herpes, human papillomavirus and HIV. The aim of this study was to find genotypic differences between the isolates. Forty isolates from prisoner's women in Tehran province were used in this study. The random amplified polymorphic DNA (RAPD) technique was used to determine genetic differ­ences among isolates and was correlated with patient's records. By each primer the banding pat­tern size of each isolates was scored (bp), genetic differences were studied, and the genealogical tree was constructed by using NTSYS software program and UPGMA method. The least number of bands were seen by using primer OPD8 and the most by using OPD3. Results showed no significant difference in isolates from different geographical areas in Iran. By using primer OPD1 specific amplified fragment with length 1300 base pair were found in only 8 isolates. All these isolates were belonged to addicted women; however, six belonged to asymptomatic patients and two to symptomatic ones. There was not much genetic diversity in T vaginalis isolates from three different geo­graphical areas.

Due to scarcity of human reports, we took advantage of the heaviest infection of M. moniliformis in rats, to describe histopathological and microanatomical valuable useful keys while confronting human occurrences. Samples were obtained from captured rats in Tehran, capital of Ira, during two dec­ades. Tissue sections were performed through hematoxylin and eosin staining to describe histopa­thological changes in rat's intestines. Totally, nine rats were found infected with M. moniliformis amongst 272 obtained rats. Heavy infection has been distinguished in 2 individuals with parasite burden of 141and 73 adult worms. Cross sections of worms within the lumen show mucosal thickness, infiltration of eosino­philic leukocyte and increase in goblet cells. Beyond the uncommonness of human infection with M. moniliformis unintended infections should not be ignored. Abundance of rats and roaches as definite and intermediate hosts must be considered particularly in countries with poor hygiene.

Southeast of Iran is an endemic area for Malaria and Crimean-Congo hemorrhagic fever (CCHF). In 1999, we faced with an outbreak of CCHF in Sistan and Baluchistan Province, in the border of Paki­stan and Afghanistan. The most cases of Malaria in Iran are also reported from this area. This arti­cle presents a 17-year- old woman who admitted to our hospital because of acute fever, head­ache, epistaxis, hemorrhagic lesions on the skin and vaginal bleeding. Finally, she was recog­nized as a case that was co -infected with CCHF and malaria.