Investigation of the antimicrobial activity of a short cationic peptide against promastigote and amastigote forms of Leishmania major (MHRO/IR/75/ER): An in vitro study.

Cutaneous leishmaniasis is one of the most endemic global health problems in many countries all around the world. Pentavalent antimonial drugs constitute the first line of leishmaniasis treatment; however, resistance to these drugs is a serious problem. Therefore, new therapies with new modes of action are urgently needed. In the current study, we examined antimicrobial activity of CM11 hybrid peptide (WKLFKKILKVL-NH2) against promastigote and amastigote forms of L. major (MHRO/IR/75/ER). In vitro anti-leishmanial activity was identified against L. major by parasite viability and metabolic activity after exposure to different peptide concentration. In the presentt study, we demostrated that different concentrations of CM11 result in dose dependent growth inhibition of Leishmania promastigotes. Furthermore, we demostrated that CM11 peptide has significant anti-leishmanial activities on amastigotes. Our results demonstrated that CM11 antimicrobial peptide may provide an alternative therapeutic approach for L. major treatment.

Identification and characterization of a cathepsin L-like cysteine protease from Rhipicephalus (Boophilus) annulatus.

The tick Rhipicephalus (Boophilus) annulatus is one of the most important ectoparasites of bovines and is responsible for the transmission of different pathogens such as Babesia and Anaplasma. Cysteine proteases are involved in several host-tick interactions including invasion of host tissues, immune evasion, pathogen transmission, embryogenesis and blood digestion. In this study, the gene encoding R. annulatus cathepsin L-like enzyme (RaCL1) was cloned into pTZ57R/T vector, sequenced and analyzed using bioinformatics approaches. The nucleotide length of RaCL1 was 999 bp. Bioinformatics analysis showed 332 amino acids with an approximate molecular weight of 36.3 kDa which contained a signal peptide sequence (18 amino acids), pro-region (97 amino acids) and mature enzyme (217 amino acids). Multiple sequence alignment of the RaCL1 revealed high similarity to cathepsin L-like cysteine proteases from other tick species such as Rhipicephalus (Boophilus) microplus and Amblyomma variegatum. Based on bioinformatics analyses, results of this work suggest that RaCL1 can be a suitable candidate for the development of vaccine against R. annulatus.

Molecular Detection and Genotyping of Intestinal Microsporidia from Stray Dogs in Iran.

BACKGROUND: Microsporidia as one of the most important pathogens in veterinary and agricultural settings, have emerged in immunocompromised patients in Iran. To date, different Enterocytozoon bieneusi genotypes have been identified in humans and animals, supporting the possibility of zoonotic zoonosis transmission potential. The aim of this study was to evaluate the distribution of E. bieneusi genotypes among overpopulated stray dogs in vicinity of Tehran, the capital city of Iran. METHODS: Totally, 75 stool and 75 urine samples were obtained from 75 stray dogs during the time period from Mar 2015 to Oct 2015. DNA extraction was performed on all the samples and specific fragment of small subunit ribosomal RNA gene of E. bieneusi and Encephalitozoon spp. was amplified. Furthermore, specific primers targeting the internal transcribed spacer region of E. bieneusi were applied to determine the genotype of the microorganism. RESULTS: Microsporidia was detected in 5.3% of stool samples, while none of the urine samples was positive for microsporidia species. Overall, 440 bp fragment of E. bieneusi was amplified in all the samples and there was no amplification for Encephalitozoon spp. The results of sequencing of 410 bp fragment of internal transcribed spacer region showed that all the E. bieneusi were genotype D. CONCLUSION: E. bieneusi was the most prevalent microsporidian species in the stray dogs and all the positive isolates were characterized as genotype D.

Detection of small number of Giardia in biological materials prepared from stray dogs.

Giardia lamblia is an intestinal protozoa with intermittent and low shedding especially in dogs, and the detection of Giardia is accompanied with problems such as sampling and diagnostic method. The objective of this study was to detection of Giardia in biological materials with low number of parasite using parasitological and molecular methods, and also to determine whether the examined stray dogs harbor known zoonotic genotype of Giardia. For this aim 85 fecal and duodenal samples were studied from which 1 was positive by Trichrome staining of stool, 4 were positive by staining of duodenal samples. The nested PCR analysis with primers derived from 18 SrRNA showed that the specific PCR product could be amplified in 4 stool and 4 duodenal samples. All positive samples in staining analysis were also positive in nested PCR. No amplification could be observed by nested PCR with primers derived from ß giardin gene due to the single copy of gene. Interestingly, the extracted DNA from old fixed stained Giardia positive smears could be also amplified with primers derived from 18SrRNA gene. The sequence analysis of nested PCR products showed that they belong to the genotype D. In conclusion, it is to denote that the Trichrome or Giemsa methods were not suitable for the detection of small number of this parasite in stool and the nested PCR with primers derived from 18S rRNA gene can replace the traditional methods successfully. For detection of Giardia in stool, primers derived from ß giardin will not be recommended.

Ectoparasites of Stray Dogs in Mazandaran, Gilan and Qazvin Provinces, North and Center of Iran.

BACKGROUND: The aim of the present study was to determine the prevalence of ectoparasite infestations in stray dogs in Mazandaran, Gilan and Qazvin Provinces in fall and winter in 2013(December to March). METHODS: Seventy dogs in 2013, from these Provinces were examined for ectoparasite infestation and diagnosis of them based on parasitological methods and identification keys was done. RESULTS: The rate of infestation in these areas was 100%, 68.5% and 93.3% respectively. Fleas were the most common ectoparasites on dogs in this study followed by lice, ticks, flies and mites respectively. The isolated arthropods were fleas in 77.5%, lice in 50%, ticks in 8.6%, flies in 6.8% and mites in 5.1% of infested dogs. The ectoparasite of the dogs included 4 flea species: Ctenocephalides canis (29.8%), C. felis (19.9%), Pulex iritans (2.9%) and Xenopsiella cheopis (0.7%), 1 louse species: Trichodectes canis (41.3%), 1 tick species: Rhipicephalus sanguinus (0.7%), 1 fly species: Hippobosca sp. (1.1%) and 1 mite species: Sarcoptes scabiei (3.6%). CONCLUSION: Fleas and lice were the most common ectoparasites in stray dogs of the studied area. Some ectoparasites transmit important human disease, therefore regular monitoring of them is a major concern to control the arthropods and arthropods-borne diseases.

Temporal and spatial distribution and species diversity of hard ticks (Acari: Ixodidae) in the eastern region of caspian sea.

Ticks are important parasites because of their voracious blood-feeding activity, and being as vectors for various agents of diseases in both human and livestock. This study was conducted in a monthly schedule from October 2014 to December 2015, at 45 study sites in three regions bordering the Caspian Sea, according to the topography including hillside, plain and coastal areas in collecting ticks from the body of sheep. Alfa and Beta biodiversity indices were calculated and compared between the three areas under study. In this study, 1583 out of a total of 3904 inspected sheep (40.54%), were infested with ticks, and 2410 ticks were collected, out of which 36.5, 34.8 and 28.7% were related to the coastal, plain and hillside areas, respectively. The identified tick specimens belonged to 3 genera and 9 species including Hyalomma marginatum, Hyalomma anatulicum, Hyalomma asiaticum, Hyalomma excavatum, Rhipicephalus bursa, Rhipicephalus turanicus, Rhipicephalus sanguineus, Haemaphysalis parva and Haemaphysalis punctata. With regards to the seasonal variation, the highest and lowest tick infestation was recognized in spring and winter, respectively. Monthly activity of ticks on sheep revealed that Hy. marginatum exhibited two peaks of activity in June and July in the coastal and plain regions, but in the hillside area these peaks for Rh. turanicus occurred in August. Differences in species richness has been proved significantly in Margalef index in the study area (P<0.05). It showed the varied number of species among sampling sites during the study period. In conclusion, substantial research in investigating ticks' population dynamics by calculating their biodiversity indices is necessary to show their interactions with the environment, as well as other species to establish the stability of their community.

Isolation of Small Number of Cryptosporidium parvum Oocyst Using Immunochromatography.

BACKGROUND: Cryptosporidium parvum causes severe gastroenteritis in immunocompromised human and new borne animals. The organism can be transmitted through water. Since small number of C. parvum is infectious, the aim of the present study was to develop a chromatography method for the isolation of C. parvum oocyst in samples with limited number of oocysts. METHODS: Antibody was prepared against whole antigen from C. parvum oocysts, the achieved Ab bound to the sepharose 4B and used for the isolation of oocysts. Antibody against P23 bound to the sepharose 4B, used also for the isolation of C. parvum oocyst. In comparison to these both methods, 2 traditional methods (Salt floatation and 55% sucrose floatation) were also performed. RESULTS: Both chromatography methods could bind oocysts with capacity depends on the column size. The isolated oocysts were free of bacteria. Our results showed that the traditional methods are useful for the isolation of oocysts from feces, in its smear stained with ziehl-nelsen, at least 3 oocyts are detectable in each microscopic field under 1000 X magnification. In contrast to the chromatography methods, the bacterial contamination was always observed in oocysts isolated with traditional methods. CONCLUSION: Immunochromatography could be used for the successful isolation of C. parvum oocysts from the samples containing limited number of oocysts.