Identification of zoonotic parasites isolated from stray dogs in bojnurd county located in North-East of Iran

Dog can represent as an important source of zoonotic disease and important health problem for human. They can carry dangerous parasitic diseases such as hydatidosis, toxocariasis and Coenurus cerebralis to humans and animals. This study was performed in order to determine the prevalence and intensity of zoonotic parasites among stray dogs from Bojnurd, the capital city of North Khorasan province in North West of Iran. During a program performing by Bojnurd municipal on the slow killing of stray dogs, 32 dogs from Jun 2013 till March 2015 were selected. At necropsy their alimentary canals were removed and to identify the species of helminthes, the nematodes were cleared in lactophenol and cestodes were stained using carmine acid. Intestinal protozoan parasites were detected with parasitological methods. 28 [87.5%] of 32 stray dogs infected at least with one helminth. Seven species of cestodes were isolated from examined dogs and three species of nematode were detected. Giardia sp. and Cryptosporidium sp. detected from fecal samples. This is the first study of the prevalence of intestinal zoonotic parasites in dogs in this area. It seems control of bearing stray dogs can help human health and reduction economic losses caused by stray dog's zoonotic parasites

Different morphologies of Leishmania major amastigotes with no molecular diversity in a neglected endemic area of zoonotic cutaneous leishmaniasis in Iran

Molecular diversity of Leishmania major and its morphological changes have become a controversial issue among researchers. Some aspects of polymorphic shapes of amastigotes in clinical manifestations along with molecular variation were evaluated among suspected patients of some exceptional zoonotic cutaneous leishmaniasis locations in Northern Khuzestan, Southwestern Iran. Suspected patients [n = 165] were sampled in zoonotic cutaneous leishmaniasis foci over two consecutive years during 2012- 2014. Prepared smears were stained, scaled and measured by ocular micrometer. DNA was extracted from smears; ITS-rDNA and Cytochrome b [Cyt b] markers were amplified, and PCR products were digested by BsuR1 restriction enzyme. Then the RFLP and sequencing were employed. Only L. major was identified in patients containing regular amastigotes' shapes [oval or round] with a size of 2-4 microm in each of classical wet, dry, mixed lesions. Meanwhile, irregular shapes [spindle, pear, or cigarette] were observed separately in non-classical wet lesions with more than 4 microm. Interestingly, a few amastigotes with an external flagellum were observed in some lesions. All sequenced ITS-rDNA and Cyt b genes of L. major did not show any molecular variation [chi [2] P > 0.05], including only one common haplotype [GenBank access no. EF413075]. Findings proved that unlike other endemic foci, there is not a meaningful correlation between phenotypic and genotypic features of L. major isolates. This study is considered as the first comprehensive report to incriminate morphometric shapes of L. major amastigotes, which enhances our knowledge concerning their relevance with various clinical appearances and genotypic traits

Novel identification of Leishmania major in Hemiechinus auritus and molecular detection of this parasite in Meriones libycus from an important foci of zoonotic cutaneous leishmaniasis in Iran

One of the well-known foci of zoonotic cutaneous leishmaniasis [ZCL] in Iran is Turkemen Sahara, which is located in north eastern Iran. ZCL is a disease of mammals, and humans can become infected as accidental hosts. Many researchers have argued that Rhombomys opimus is the only main reservoir host of ZCL in this region of the Golestan province. No other rodents or mammals are thought to host or have been reported to host Leishmania parasites in this region. This research was designed and developed to isolate, detect and firmly identify Leishmania parasites in mammals and rodents other than R. opimus. Wild mammals were caught from gerbil burrows. Leishmania parasites were detected to assess the infection of reservoir hosts in 2010. Each genomic DNA sample was screened for Leishmania infection via nested PCR and sequencing using the internal transcribed spacer ribosomal DNA [ITS-rDNA] identification protocol for parasites. The greatest number of infections [8/19] were found in Menones libycus. One in three infections was found in Hemiechinus auritus, and this is the first report of infection in this species. Only Leishmania major was definitively identified and unambiguously typed in M. libycus and H. auritus. The infection rates in these two wild mammals were not significantly different, and no other gerbil parasites were detected in M. libycus or H. auritus at our study site. Recent findings of Leishmania turanica in R. opimus and failures to detect L. turanica in M. libycus may be attributable to unidentified Leishmania infections in two M. libycus due to unreadable sequences. These cases may represent mixed infections by L. major and L. turanica. The assumptions that gerbil parasites can be co-infectors provide a starting point for the identification of the causative and potential parasites responsible for the frequent infections that are mainly mediated via sandfly vectors

[Scolicidal effects of Berberis vulgaris fruit extract on hydatid cyst protoscolices]

Background: Surgery is one of the best choices for the treatment of hydatidosis

The use of effective scolicidal agents during surgery for hydatid cyst is essential to prevent the secondary infection. Up to now no effective and safe agent has been identified for this purpose

Berberis vulgaris called [Zereshk] in Persian has been traditionally used as herbal remedy for the treatment of complaints and it is widely cultivated in Iran. Many studies have shown that it has antibacterial, antifungal and antiparasitic effect

Methods: In this study the scolicidal effect of Berberis vulgaris aqueous and hydro-alcohol extract for different concentrations [for aqueous: 0.5, 1, 2 and 4 nig/ml and for hydro-alcohol: 0.25, 0.5, 1 and 2 mg/ml] at different exposure times [5, 15 and 30 minutes] was evaluated. For this purpose, we obtained liver hydatid-cysts from a slaughter house

Viability of protoscolices was assessed by 0.1% eosin staining. Normal saline and hypertonic saline were used as negative and positive controls respectively

Results: All the different concentrations of Berberis vulgaris aqueous and hydro-alcoholic extracts had scolicidal effect

An aqueous extract with 4mg/ml concentration acted as positive control and we observed to lead to the death of 100% of protoscolices in the first 5 minutes

The least scolicidal effect [12.3%] was observed in an aqueous extract with 0.5 mg/ml concentration.The scolicidal activity of hydro-alcoholic extract with concentration of 2 mg/ml was 100% after 5 min of application, which was the same as positive control group

We noticed a significant increase in protoscolicidal activity with an increase in concentration in the two extracts of Berberis vulgaris [P<0.001]

Conclusion: It is important to mention that all the concentration levels and exposure times applied in this experiment are relatively low, since scolitical activity in both of the extracts is at its highest in this low spectrum. For further experiments, we suggest that the stability of cyst fluid in both of the extracts should be assessed

Therefore, after In vivo examination and additional experiments, it may be used as a suitable and effective scolicidal in surgery

Genotyping and phylogenetic analysis of fasciola spp. isolated from sheep and cattle using PCR-RFLP in Ardabil province, Northwestern Iran

The aim of this study was to detect the genotype of Fasciola spp. in Meshkin-Shahr, Ardabil Province, northwestern Iran in different hosts using PCR-RFLP. The parasite hosts included cattle, and sheep. Overall, 70 adult flukes from livers of slaughtered animals were collected from the abattoirs of aforementioned area. The included 35 samples from infected sheep and 35 samples from 35 infected cattle. PCR-RFLP and sequence analysis of the first nuclear ribosomal internal transcribed spacer [ITS 1] region from Fasciola species were used to conduct the study. The fragment of approximately 700bp in all of the Fasciola samples was amplified. PCR products of ITS 1 were subjected for digestion by restriction enzyme. RsaI restriction enzyme was selected for RFLP method that caused the separation specifically of Fasciola species. Amplicons with the sequences of F. hepatica had a pattern of about 360, 100, and 60 bp band size, whereas F. gigantica worms had a profile of 360, 170, and 60 bp in size, respectively. Results based on PCR-RFLP analysis were confirmed by sequence analysis of representative ITS 1 amplicons. No hybrid forms were detected in the present study. All sheep were infected with F. hepatica but cattle were infected with both species. Both species of Fasciola are present in Ardabil. The method described here can be valuable for identification of Fasciola species in endemic parts for fasciolosis, regions with intermediate species and in that overlapping distribution area

Identification of infective larva [L3] proteins of Strongyloides stercoralis by immunoblot

Strongyloides stercoralis is prevalent in tropical and subtropical regions worldwide. This parasite is the only nematode with the ability to multiply in its host's body via autoinfection transmission. Larvae detection in feces is difficult partly because of low egg production and also irregular larvae excretion in feces. Serologic tests [ELISA, IFA] are also diagnostic, however Strongyloides stercoralis antigens are not available as a diagnostic tool. In the present study, we analyzed filariform larva [L3] proteins of Strongyloides stercoralis by the immunoblot technique. Stool samples were examined by direct smear, formalin-ether and agar plate method to identify infected patients. Sera were also obtained and stored at -20°C. Infective larvae were then obtained by agar plate culture, which was incubated for 6-7days at 20°C, then frozen at -70°C. Finally, larvae were suspended at a concentration level of 12000 in 250ml PBS, containing protease inhibitors and then were sonicated. Protein level was measured by Bradford method. Proteins of Strongyloides stercoralis filariform larvae were separated by SDS-PAGE, blotted onto nitrocellulose paper. Western blot analysis of these antigens was achieved using infected human sera [0.1, 0.01, 0.001 dilution] with strongyloidiasis, toxocariasis, hydatidosis, amebiasis and normal human serum as control. Four immunodominant proteins [23, 28, 30, 41 kDa] were recognized with strongyloidiasis sera in 0.1 diluted serum. None of the proteins reacted to normal human and amebiasis serum, but some showed reaction with serum of hydatidosis and toxocariasis. Having increased the level of serum dilution, only 41 kDa protein was recognized by strongyloidiasis sera. Other serums did not represent any reaction to the parasite's proteins. Therefore, the 41 kDa protein presents as the most important immunodominant protein in this study. The identification of immunodominant proteins adapted to the physiological and genetic conditions of the host is an appropriate diagnostic approach, which could be associated with improved sensitivity and specificity of serologic tests