Contamination of tap water sources to Acanthamoeba spp. in selected cities of Hormozgan province, Iran

Background    Free-living amoeba (FLA) such as Acanthamoeba are ubiquitous and amphizoic protozoan parasites. These parasites can cause severe human infections. Water resources are one of the important habitats for FLAs. Waters in tanks and wells can be a habitat for Acanthamoeba species, particularly in warm climates. The present study was conducted to evaluate the presence of Acanthamoeba species in tap water resources of selected cities of Hormozgan Province, Southern Iran.Material and methods In this cross sectional study, 50 water samples (approximately 500 mL) were collected from tap water resources (25 samples from tanks and 25 samples from wells) of Hormozgan province. The samples were filtered, cultured on 1.5% non-nutrient agar medium. After incubation at room temperature, cloning was performed. Investigation of Acanthamoeba species was performed using the microscopic method and page key.Results Out of 50 samples, 14 samples (28%) were found positive. Out of 25 examined well samples 9 samples (36%) were found contaminated with Acanthamoeba spp., while only 5 (20%) of 25 tank samples were found positive. All samples were cloned successfully.Conclusion The high occurrence of Acanthamoeba spp. in water resources of Hormozgan province could be as a hazard for public health of the region, and using effective disinfectants and accurate monitoring may be a good suggestion. 

A survey of Free-Living Amoebae (FLAs) in Nasal Mucosa of Healthy Individuals using Culture-based Method

Free-living amoebas (FLAs) are facultative parasites, and some of them can be highly pathogenic for animals and humans. These protozoan are found in abundance in environmental resources such as freshwater and soil, and thus the possible colonization of such FLA is probable. Our previous studies showed that Acanthamoeba spp. can colonize nasal mucosa of immunosuppressed patients. However, there was no report of colonization of free-living amoebae in healthy individuals in Iran.  The present study was conducted to investigate the possibility that FLAs are the normal flora of the nasal mucosa. Overall, 46 nasal swab samples were collected from healthy individuals and cultured on a 1.5% non-nutrient agar (NNA) plate enriched with a lawn of heat-killed Escherichia coli. The plates were incubated at room temperature. After one week, the plates were microscopically examined, and this was continued for one month. None of the samples were found positive for FLAs. Our findings may indicate that FLAs are not the normal flora of the nasal mucosa, or their number is so limited in the nasal mucosa that they cannot be isolated by the method used. However, supplementary and comprehensive studies are recommended

Comparison of Three Commonly Used Genetic Markers for Detection of Leishmania Major: An Experimental Study

BACKGROUND፡ Leishmaniasis is a vector-borne disease caused by an intracellular protozoan parasite called Leishmania spp. Different species produce different clinical outcomes; the majority of cases are cutaneous forms. Leishmania major is one of the main causative agents of cutaneous leishmaniasis (CL). Various methods are being using to diagnose CL, including microscopic examination, culture, and molecular detection of the parasite genome.METHOD: In the current study, we tried to compare three common molecular markers, including Kinetoplast DNA (kDNA), Cytochrome b (Cyt b), and Internal transcribed space 1 (ITS1), for the detection of Leishmania major. After cultivation of standard strain of L. major MHOM/IR/75/ER in RPMI 1640, certain number of promastigotes was subjected to DNA extraction and different PCR reactions.RESULTS: The lowest number of the parasite (5 promastigotes) can be detected by kDNA-PCR, followed by Cyt b-PCR (10 promastigotes), and ITS1-PCR (50 promastigotes).CONCLUSION: In conclusion, kDNA-PCR was the most sensitive marker and may provide more reliable data in the initial screening, especially in false-negative results provided by parasitological methods due to the low number of parasites

Molecular Characterization of Pathogenic Acanthamoeba Isolated from Drinking and Recreational water in East Azerbaijan, Northwest Iran

Acanthamoeba -related infections, such as amoebic keratitis and granulomatous amoebic encephalitis, can develop in high-risk population through contaminated water sources. Thus, surveying water resources, particularly those available for human use, is of the utmost importance. In the present study, 67 water samples were collected from water resources in East Azerbaijan, a province in northwestern Iran. Samples were cultured on enriched non-nutrient agar plates, and sequencing-based approaches were used for genotyping. The pathogenic potential of the isolates was determined using thermo- and osmo-tolerance tests. Acanthamoeba were detected in 17 (25.4%) of the 67 collected samples. Sequencing analysis revealed that the isolates belonged to the T3 (23.52%), mixed T3/T4 (5.88%), T4 (58.82%), T5 (5.88%), and T13 (5.88%) genotypes. Through thermo- and osmo-tolerance tests, 88.23% of isolates were resistant to 37 °C, 40 °C temperature, and 0.5 M and 1 M osmolarity; thus, these isolates had the potential for pathogenicity. These findings point toa serious public health concern in the studied region. This study is the first to report Acanthamoeba isolated from drinking and recreational water sources in East Azerbaijan and Acanthamoeba T13 isolated from tap water in Iran

Morphological and Molecular Identification of Acanthamoeba Spp from Surface Waters in Birjand, Iran, During 2014-2015

Background & Aims of the Study: Free-living amoebae (FLA) are opportunistic and ubiquitous protozoa that are widely found in various environmental sources. They are known to cause serious human infections including a fatal encephalitis, a blinding keratitis, and pneumonia. So, due to their medical importance, the identification of free living amoeba in water resources, as a source of human infection, is necessary. The objective of this study was to isolate the Acanthamoebaspp from the surface waters of Birjand, Iran, during 2014-2015 by Morphological and molecular method. Materials and Methods:  In a cross-sectional study, 50 samples were collected from different localities of Birjand city including the surface waters, pools and fountains in parks,squares and water stations from the October 2014 to the January 2015.Each sample was filtered through a nitrocellulose membrane filters and cultured on non-nutrient agar (NNA) with Escherichia coli suspension and incubated for 1 week to 2 months at room temperature.The plates were examined by the microscopy to morphologically identify Acanthamoeba species. Following DNA extraction, PCR specific primers was used to confirm the identification morphologically. Results:  Out of 50 water samples, 19 (38%) were positive for Acanthamoebatrophozoites and cysts according to the morphological criteria. In addition, Acanthamoebaspp was identified by PCR method, using genus specific primers pairs in 15 (78.9%) cases of positive cultures, showing anearly 500bp band. Conclusion: According to the prevalent of Acanthamoebaspp in the surface stagnant waters of Birjand, more attention to the potential role of such waters in transmission of infection by the regional clinicians and health practitioners is necessary

Genotyping of Toxoplasma gondii Strains Isolated from Patients with Ocular Toxoplasmosis in Iran

Background: Toxoplasma gondii is an obligate, intracellular protozoon that develops its sexual stage in cat’s intestinal epithelial cells as definitive host and develops its asexual stage in different tissues of a wide range hosts called intermediate host. The protozoon is a food-borne and worldwide parasite that can cause serious complications such as abortion in pregnant women, encephalitis, and ocular toxoplasmosis. The present study aimed to genotype T. gondii strains isolated from patients with toxoplasmic retinochoroiditis. Methods: Fifty-two blood samples were taken from patients with ocular toxoplasmosis, from July 2013 to July 2014. The specimens were collected from three ophthalmological hospitals of Tehran, Iran. After that, DNA extraction was performed using kit on separated buffy coats of serologically positive blood samples. Then PCR was done in GRA6 gene. For digestion of products, MseI endonuclease was used. Finally, some of the PCR products were sequenced. Results: All of 52 samples were found positive by serological and PCR-RFLP methods and all of isolated strains belong to type III genotype. Type III genotype has the highest prevalence in Iranian ocular toxoplasmic patients. Conclusion: T. gondii, particularly its type III should not be neglected as a cause of retinochoroiditis

Microscopic isolation and characterization of free living amoebae (FLA) from surface water sources.in Birjand, the capital city of the South Khorasan

Background and Aim: Free living amoebae (FLA) are amphizoic protozoa that are widely found in various environmental sources. They are known to cause serious infections in human and animal nervous systems. . The aim of the current study was to determine the presence of Acanthamoeba spp in surface water sources in Birjand city employing microscopic culture analysis based on morphology features of the amoebae. Materials and Methods: In this cross-sectional study, 50 samples of surface water sources in Birjand city. including . parks pools, public squares, waterfronts, and water stations were collected and .transferred to the laboratory and were passed through nitrocellulose filter paper. The remained elements in the filters were cultured on non-nutrient agar (NNA) with 100μl Escherichia coli suspension. After a few weeks of using morphological features,the amoeba grown were identified. Results: Out of the total of 50 samples cultured on non-nutrient agar (NNA), 19 (38%) samples.were morphologically polluted with Acanthamoeba spp,. In 2 samples (4%) a colony of Vahlkampfiidae were observed. Conclusion: The results indicated that a significant percentage of surface water sources in Birjand city was contaminated with Acanthamoeba spp. It is necessary for physicians, therefore, to take into account the diseases caused by these infectious agents. Besides, local regional health professionals should take into consideration the potential role of surface stagnant water sources in transferring these infectious agents. Placing warning signs in areas contaminated with these infectious agents seems a useful measure

Morphological and Molecular Survey of Naegleria spp. in Water Bodies Used for Recreational Purposes in Rasht city, Northern Iran

Background: Naegleria spp. is a free-living amoeba of which some species includ­ing N. fowleri and N. australeinsis are highly pathogenic in human and ani­mals. These widespread amoebae could be found in different environmental sources particularly in aquatic resources of tropical and subtropical regions. The most important source of infection is via recreational water contact. Due to the lack of thorough research regarding species of Naegleria spp. in aquatic sources, the present study was conducted. Methods: In the present study, 60 samples were collected from recreational wa­ter resources of Rasht city, Guilan province, north of Iran. After filtering and culturing the samples, plates were examined by microscopic method and accord­ing to the page criteria. DNA of vahlkampfiid-positive samples were then ex­tracted using phe­nol-chlorophorm method. Amoebae genus was identified by targeting the ITS-region and sequencing based-approaches. Results: Nine (15%) samples out of a 60 total samples were positive for Naegleria spp. of which seven belonged to potentially pathogenic N. australiensis. Two other strains were belonged to non-pathogenic N. pagei. Conclusion: The present research was the first report of occurrence of N. aus­traliensis and N. pagei in Rasht city, north Iran. This study reflects the occurrence of Naegleria spp. in water sources of Guilan Province, Iran