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Heliyon Sep 2020causes severe diseases such as Granulomatous Amebic Encephalitis (GAE) and keratitis (AK). Improving the culture media classically used for this amoeba could help to...
causes severe diseases such as Granulomatous Amebic Encephalitis (GAE) and keratitis (AK). Improving the culture media classically used for this amoeba could help to identify it quickly and facilitate its study as a biological model. The purpose of this study was to compare the growth of two genotypes (T3 and T4) in several culture media. (T3 genotype) and (T4 genotype) were cultured in PYG, TSY, TYI-S-33, RPMI, and RPMI-FBS medium. The number of amoebas grown in different culture media was counted and compared to each other for 14 days. Findings in this research revealed the highest growth in RPMI-FBS medium. For this reason, we can recommend this culture medium to promote the growth of in its biological studies.
PubMed: 32984575
DOI: 10.1016/j.heliyon.2020.e04805 -
Genome Biology and Evolution Oct 2020Peroxisomes perform various metabolic processes that are primarily related to the elimination of reactive oxygen species and oxidative lipid metabolism. These organelles...
Ultrastructural, Cytochemical, and Comparative Genomic Evidence of Peroxisomes in Three Genera of Pathogenic Free-Living Amoebae, Including the First Morphological Data for the Presence of This Organelle in Heteroloboseans.
Peroxisomes perform various metabolic processes that are primarily related to the elimination of reactive oxygen species and oxidative lipid metabolism. These organelles are present in all major eukaryotic lineages, nevertheless, information regarding the presence of peroxisomes in opportunistic parasitic protozoa is scarce and in many cases it is still unknown whether these organisms have peroxisomes at all. Here, we performed ultrastructural, cytochemical, and bioinformatic studies to investigate the presence of peroxisomes in three genera of free-living amoebae from two different taxonomic groups that are known to cause fatal infections in humans. By transmission electron microscopy, round structures with a granular content limited by a single membrane were observed in Acanthamoeba castellanii, Acanthamoeba griffini, Acanthamoeba polyphaga, Acanthamoeba royreba, Balamuthia mandrillaris (Amoebozoa), and Naegleria fowleri (Heterolobosea). Further confirmation for the presence of peroxisomes was obtained by treating trophozoites in situ with diaminobenzidine and hydrogen peroxide, which showed positive reaction products for the presence of catalase. We then performed comparative genomic analyses to identify predicted peroxin homologues in these organisms. Our results demonstrate that a complete set of peroxins-which are essential for peroxisome biogenesis, proliferation, and protein import-are present in all of these amoebae. Likewise, our in silico analyses allowed us to identify a complete set of peroxins in Naegleria lovaniensis and three novel peroxin homologues in Naegleria gruberi. Thus, our results indicate that peroxisomes are present in these three genera of free-living amoebae and that they have a similar peroxin complement despite belonging to different evolutionary lineages.
Topics: Acanthamoeba castellanii; Balamuthia mandrillaris; Catalase; Microscopy, Electron, Transmission; Peroxins; Peroxisomes; Phylogeny
PubMed: 32602891
DOI: 10.1093/gbe/evaa129 -
Parasite Immunology Mar 2020The aim of this study was to evaluate the inflammation process that resulted from the inoculation of Wistar Rats with Acanthamoeba griffini, a virulent T3 Acanthamoeba...
The aim of this study was to evaluate the inflammation process that resulted from the inoculation of Wistar Rats with Acanthamoeba griffini, a virulent T3 Acanthamoeba genotype that produces keratitis. Haematoxylin and eosin, periodic acid stain, immunohistochemistry and morphometry were used to analyse tissues from rats of an Acanthamoeba keratitis (AK) model. Two weeks after inoculating the rats with A griffini trophozoites, the thickness of the stroma had diminished, followed by an increase in thickness at 4 weeks. At the latter time, an abundance of inflammatory infiltrate cells was observed, some found to express IL-1β, IL-10 and/or caspase 3. Intercellular adhesion molecule-1 was expressed in corneal blood vessels amid the abundant vascularization characteristic of the development of AK. Through an immunohistochemical technique, trophozoites were detected at 2 and 4 weeks post-inoculation. By 8 weeks, there were a low number of trophozoites and cysts and the corneas of infected rats were similar in thickness to those of the controls. Thus, the rats were capable of healing experimental AK in the present rat model. Diverse immunological mechanisms regulated the inflammatory process in acute AK induced by A griffini in a murine model.
Topics: Acanthamoeba; Acanthamoeba Keratitis; Animals; Apoptosis; Caspase 3; Cornea; Disease Models, Animal; Female; Humans; Intercellular Adhesion Molecule-1; Interleukin-10; Interleukin-1beta; Mice; Rats; Rats, Wistar; Trophozoites
PubMed: 31856305
DOI: 10.1111/pim.12692 -
Genotyping determination of Acanthamoeba strains: an original study and a systematic review in Iran.Journal of Water and Health Oct 2019This study aimed to detect the presence of Acanthamoeba spp. in different water resources of Zahedan, southeast of Iran, and also systematically reviewed all...
This study aimed to detect the presence of Acanthamoeba spp. in different water resources of Zahedan, southeast of Iran, and also systematically reviewed all publications regarding Acanthamoeba in Iran (2005-2018). Fifty water samples were collected from different water resources in Zahedan. The positive samples were identified morphologically and subjected to polymerase chain reaction (PCR) using fragments of 18S rRNA. In the systematic review, data collection using particular terms was carried out using the following electronic databases including Science Direct, ISI Web of Science, MEDLINE, EBSCO, Scopus, and Google Scholar. A total of 17 (34%) samples were positive for Acanthamoeba spp., and nucleotide sequencing indicated that 15 samples (88.23%) belonged to the T4 genotype and the rest belonged to the T5 genotype. A total of 39 studies reported genotyping of Acanthamoeba spp. from various geographical areas of Iran and revealed that T4 (35 studies), T5 (19 studies), T3 (11 studies), T11 (8 studies), and T2 (6 studies) genotypes were the most prevalent in Iran. The T4 genotype of Acanthamoeba is a prevalent free-living amoeba and widely distributed not only in Zahedan but also in other provinces of Iran. Phylogenetic analysis reveals that A. castellanii and A. griffini predominantly colocalize with the T4 genotype.
Topics: Acanthamoeba; Environmental Monitoring; Fresh Water; Genotype; Iran; Phylogeny; RNA, Ribosomal, 18S; Water Supply
PubMed: 31638023
DOI: 10.2166/wh.2019.048 -
Parasitology Research Jun 2019The genus Acanthamoeba can cause Acanthamoeba keratitis (AK) and granulomatous amoebic encephalitis (GAE). The treatment of these illnesses is hampered by the existence...
The genus Acanthamoeba can cause Acanthamoeba keratitis (AK) and granulomatous amoebic encephalitis (GAE). The treatment of these illnesses is hampered by the existence of a resistance stage that many times causes infection relapses. In an attempt to add new agents to our chemotherapeutic arsenal against acanthamebiasis, two Acanthamoeba isolates were treated in vitro with newly synthesized biguanide dendrimers. Trophozoite viability analysis and ultrastructural studies showed that dendrimers prevent encystment by lysing the cellular membrane of the amoeba. Moreover, one of the dendrimers showed low toxicity when tested on mammalian cell cultures, which suggest that it might be eventually used as an amoebicidal drug or as a disinfection compound in contact lens solutions.
Topics: Acanthamoeba; Acanthamoeba Keratitis; Amebicides; Animals; Biguanides; Cell Line, Tumor; Chlorhexidine; Contact Lens Solutions; Dendrimers; Encephalitis; HeLa Cells; Humans; Trophozoites
PubMed: 31069536
DOI: 10.1007/s00436-019-06341-7 -
European Journal of Protistology Apr 2019Nuclear group I introns are parasitic mobile genetic elements occurring in the ribosomal RNA genes of a large variety of microbial eukaryotes. In Acanthamoeba, group I...
Nuclear group I introns are parasitic mobile genetic elements occurring in the ribosomal RNA genes of a large variety of microbial eukaryotes. In Acanthamoeba, group I introns were found occurring in the 18S rDNA at four distinct insertion sites. Introns are present as single elements in various strains belonging to four genotypes, T3 (A. griffini), T4 (A. castellanii complex), T5 (A. lenticulata) and T15 (A. jacobsi). While multiple introns can frequently be found in the rDNA of several algae, fungi and slime moulds, they are usually rare and present as single elements in amoebae. We reported herein the characterization of an A. lenticulata strain containing two introns in its 18S rDNA. They are located to already known sites and show basal relationships with respective homologous introns present in the other T5 strains. This is the first and unique reported case of multiple nuclear introns in Acanthamoeba.
Topics: Acanthamoeba; DNA, Protozoan; Introns; RNA, Ribosomal, 18S; Species Specificity
PubMed: 30743186
DOI: 10.1016/j.ejop.2019.01.007 -
Scientific Reports Sep 2018Gemmata spp. bacteria thrive in the same aquatic environments as free-living amoebae. DNA-based detection of Gemmata spp. sequences in the microbiota of the human...
Gemmata spp. bacteria thrive in the same aquatic environments as free-living amoebae. DNA-based detection of Gemmata spp. sequences in the microbiota of the human digestive tract and blood further questioned the susceptibility of Gemmata spp. to phagocytes. Here, Gemmata obscuriglobus and Gemmata massiliana were co-cultured with the amoebae Acanthamoeba polyphaga, Acanthamoeba castellanii, Acanthamoeba griffini and THP-1 macrophage-like phagocytes. All experiments were performed in five independant replicates. The ratio amoeba/bacteria was 1:20 and the ratio THP-1/bacteria was 1:10. After a 2-hour co-culture, extracellular bacteria were killed by kanamycin or amikacin and eliminated. The intracellular location of Gemmata bacteria was specified by confocal microscopy. Microscopic enumerations and culture-based enumerations of colony-forming units were performed at T = 0, 1, 2, 3, 4, 8, 16, 24, 48 and 72 hours post-infection. Then, Gemmata bacteria were engulfed into the phagocytes' cytoplasmic vacuoles, more than (98 ± 2)% of Gemmata bacteria, compared to controls, were destroyed by phagocytic cells after a 48-h co-culture according to microscopy and culture results, and no positive culture was observed at T = 72-hours. Under our co-culture conditions, Gemmata bacteria were therefore susceptible to the environmental and host phagocytes here investigated. These data suggest that these Acanthamoeba species and THP-1 cells cannot be used to isolate G. massiliana and G. obscuriglobus under the co-culture conditions applied in this study. Although the THP-1 response can point towards potential responses that might occur in vivo, these responses should first bevalidated by in vivo studies to draw definite conclusions.
Topics: Acanthamoeba; Coculture Techniques; Humans; Macrophages; Planctomycetales; THP-1 Cells
PubMed: 30190504
DOI: 10.1038/s41598-018-31667-0 -
Mikrobiyoloji Bulteni Jul 2018The free living amoebae cause various infections such as Acanthamoeba keratitis, granulomatous amoebic encephalitis, primer amoebic meningoencephalitis in humans and...
The free living amoebae cause various infections such as Acanthamoeba keratitis, granulomatous amoebic encephalitis, primer amoebic meningoencephalitis in humans and animals. The free living amoebae Acanthamoeba, Balamuthia mandriallis, Naegleria fowleria and Sappinia species that cause disease in humans have been isolated from many environmental materials until today. However, no isolation has been reported from the filters of the air conditions from the houses used for ventilation. The aim of this study was toinvestigate the existence of free living amoebae using molecular methods in the filters of air-conditions used in the study living area of the people. A total of 30 dust samples were taken from the filtersof air-conditions in Adana and Gaziantep province of Turkey. DNA isolation of the dust samples was performed using the DNeasy PowerSoil kit (Qiagen, Germany) and polymerase chain reaction was done with specific primers of Acanthamoeba spp., B.mandriallis, N.fowleria and Sappinia species. As a result of this study, Acanthamoeba spp. was determined as 33.3% (5/15) and B.mandriallis was determined as6.6% (1/15) in Adana province. On the other hand, Acanthamoeba species was determined as 26.6% (4/15) and B.mandriallis was determined as 13.3% (2/15) in Gaziantep province. N.fowleria and Sappina species were not detectedin both of the cities. DNA sequence analysis was performed for the confirmation of the species and 99% of the results were similar to the other species in GenBank. The rates of Acanthamoeba castellanii and Acanthamoeba griffinii (T3) were determined as %66.6 (6/9) and 33.3% (3/9), respectively by DNA sequencing. Distribution of Acanthamoeba species according to the cities were 33.3% (3/9) for A.castellanii and 22.2% (2/9) for A.griffini in Adana. It was 33.3% (3/9) for A.castellanii and 11.1% (1/9) for A.griffini in Gaziantep. There was no significant difference in the distribution of the parasite species among cities (p> 0.1). It is important to raise awareness of the diseases caused by free living amoebae among people. Acanthamoeba species have been reported frequently from environmental materials in Turkey, but B.mandriallis has not been reported from any environmental sample since this study. The presence of B.mandriallis has been reported in the air-conditions of houses in this study. This result shows that people have risk in terms of illness of free living amoebae in living areas. Our study emphasized that firstly the health personnel and then the people should be informed about the deadly parasites and the cleaning of the air conditions should be done in certain periods.
Topics: Air Conditioning; Amebiasis; Balamuthia mandrillaris; Household Articles; Humans; Turkey
PubMed: 30156514
DOI: 10.5578/mb.67092 -
European Journal of Protistology Oct 2018Various strains belonging to three Acanthamoeba species, A. griffini (genotype T3), A. lenticulata (T5), and A. jacobsi (T15), have group I introns in their 18S rRNA...
Various strains belonging to three Acanthamoeba species, A. griffini (genotype T3), A. lenticulata (T5), and A. jacobsi (T15), have group I introns in their 18S rRNA genes. Group I introns are self-splicing ribozymes that can spread among host lineages either through an intron-encoded endonuclease at the DNA level, or by reverse splicing during the RNA cycle. In Acanthamoeba, introns belong to the subclass IC1, they are located at one out four positions within the rRNA, show low identity values and all lack open reading frames to encode for an endonuclease. Uncharacterized introns from strains of another genotype, T4 (A. castellanii complex), resemble those of genotype T3, and at least one of them contains a non-functional endonuclease gene. Here, we analyzed all available data on Acanthamoeba 18S rDNA sequences to identify the possible presence of open reading frames that could encode endonucleases. We found a total of eight 18S rDNA sequences, all from T4 strains, that have introns containing putative non-functional endonuclease genes. Furthermore, two distinct endonucleases can be identified that are differently inserted in unrelated introns.
Topics: Acanthamoeba; Endonucleases; Genotype; Introns; RNA, Ribosomal, 18S; Species Specificity
PubMed: 30071371
DOI: 10.1016/j.ejop.2018.07.002 -
Microbial Pathogenesis Jan 2018Mycobacterium ulcerans, a decaying Mycobacterium marinum derivative is responsible for Buruli ulcer, a notifiable non-contagious disabling infection highly prevalent in...
Mycobacterium ulcerans, a decaying Mycobacterium marinum derivative is responsible for Buruli ulcer, a notifiable non-contagious disabling infection highly prevalent in some West African countries. Aquatic environments are suspected to host M. ulcerans, however, the exact reservoirs remain unknown. While M. marinum was found to resist amoebal microbicidal activities, this remains unknown for M. ulcerans. In this study M. ulcerans was co-cultured with the moderately halophile Acanthamoeba griffini at 30 °C to probe this tropical amoeba as a potential reservoir for M. ulcerans. In triplicate experiments, we observed engulfment of M. ulcerans by A. griffini trophozoites, followed by an unexpected significant difference of 98.4% (day 1), 99.5% (day 2), 99.5% (day 3) and 99.9% (day 7) between the number of intra-amoebal mycobacteria detected by PCR and the number of viable intra-amoebal mycobacteria measured by 10-week culture. Further encystment revealed only one Mycobacterium organism for 150 A. griffini cysts observed by electron microscopy and the culture of excysted amoebae remained sterile. In conclusion, these data install M. ulcerans as susceptible to A. griffini microbicidal activities rendering this amoeba species an unlikely host of M. ulcerans in natural environments.
Topics: Acanthamoeba; Amoeba; Buruli Ulcer; Coculture Techniques; DNA, Bacterial; Disease Reservoirs; Environmental Microbiology; Microbial Viability; Mycobacterium ulcerans
PubMed: 29155010
DOI: 10.1016/j.micpath.2017.11.022