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Frontiers in Cellular and Infection... 2023Entamoeba is an anaerobic protozoan that is strongly associated with inflamed periodontal pockets. It is able to invade the mucosal epithelium of the human host, where...
BACKGROUND
Entamoeba is an anaerobic protozoan that is strongly associated with inflamed periodontal pockets. It is able to invade the mucosal epithelium of the human host, where it can feed on epithelial cells and elicit a severe innate immune response. Unlike other species, it is considered that cannot form cysts, because it is a non-infectious protozoan. The lack of encystation capability would make it susceptible to periodontal treatment. However, it is not clear how the human host becomes infected with trophozoites. We investigated the ability of to encapsulate in response to an unfavorable environment .
METHODS
Different strains of , isolated from inflamed periodontal pocket samples, were cultured for 8 days in the presence or absence of the antimicrobials amoxycillin and metronidazole. To reveal cyst formation, we investigated the morphology and ultrastructure of the amoeba by light, fluorescence, transmission and scanning electron microscopy. We also used the fluorescent dye calcofluor white M2R to demonstrate chitin present in the cyst wall.
RESULTS
We observed exocysts and an intra-cystic space separating the encapsulated trophozoite from the environment. Remarkably, cysts showed a smooth surface, polygonal edges and smaller size compared to free-living trophozoites. In addition, encapsulated trophozoites that detached from the cyst wall had a dense cytoplasma without phagocytic vesicles. The cyst walls consisted of chitin as in other species. The encapsulated trophozoids were mononuclear after antibioticinduced encapsulation.
DISCUSSION
We conclude that cyst formation has significant implications for dissemination and infection and may explain why established treatment approaches often fail to halt periodontal tissue destruction during periodontitis and peri-implantitis.
Topics: Animals; Humans; Entamoeba; Trophozoites; Cysts; Anti-Bacterial Agents; Chitin
PubMed: 37469604
DOI: 10.3389/fcimb.2023.1201394 -
Systematic and Applied Microbiology Nov 2023Chlamydiota are an ancient and hyperdiverse phylum of obligate intracellular bacteria. The best characterized representatives are pathogens or parasites of mammals, but...
Chlamydiota are an ancient and hyperdiverse phylum of obligate intracellular bacteria. The best characterized representatives are pathogens or parasites of mammals, but it is thought that their most common hosts are microeukaryotes like Amoebozoa. The diversity in taxonomy, evolution, and function of non-pathogenic Chlamydiota are slowly being described. Here we use data mining techniques and genomic analysis to extend our current knowledge of Chlamydiota diversity and its hosts, in particular the Order Parachlamydiales. We extract one Rhabdochlamydiaceae and three Simkaniaceae Metagenome-Assembled Genomes (MAGs) from NCBI Short Read Archive deposits of ciliate and algal genome sequencing projects. We then use these to identify a further 14 and 8 MAGs respectively amongst existing, unidentified environmental assemblies. From these data we identify two novel clades with host associated data, for which we propose the names "Sacchlamyda saccharinae" (Family Rhabdochlamydiaceae) and "Amphrikana amoebophyrae" (Family Simkaniaceae), as well as a third new clade of environmental MAGs "Acheromyda pituitae" (Family Rhabdochlamydiaceae). The extent of uncharacterized diversity within the Rhabdochlamydiaceae and Simkaniaceae is indicated by 16 of the 22 MAGs being evolutionarily distant from currently characterised genera. Within our limited data, there was great predicted diversity in Parachlamydiales metabolism and evolution, including the potential for metabolic and defensive symbioses as well as pathogenicity. These data provide an imperative to link genomic diversity in metagenomics data to their associated eukaryotic host, and to develop onward understanding of the functional significance of symbiosis with this hyperdiverse clade.
Topics: Animals; Metagenome; Chlamydiales; Phylogeny; RNA, Ribosomal, 16S; Bacteria; Metagenomics; Mammals
PubMed: 37847957
DOI: 10.1016/j.syapm.2023.126468 -
Development and application of recombinase polymerase amplification assay for rapid detection of sp.Parasitology Nov 2023sp. is a common parasite in the intestinal tract of humans and animals. The clinical diagnosis of sp. mainly depends on the microscopic observation of parasite, which...
sp. is a common parasite in the intestinal tract of humans and animals. The clinical diagnosis of sp. mainly depends on the microscopic observation of parasite, which can lead to false-negative results. An accurate and convenient diagnostic approach for sp. infection is crucial for effectively preventing and controlling blastocystosis. Herein, we developed a recombinase polymerase amplification (RPA) method for detecting sp. The results showed that the DNA amplification by RPA established in this study could be performed within 5 min at 37°C, with maximum band intensity observed at 30 min. The minimum detection limit of RPA was 100 fg L, consistent with conventional polymerase chain reaction (cPCR). Furthermore, the RPA method exhibited no cross-reactivity with 7 other non-target pathogens in the intestinal tract. Next, the newly established RPA method was used to analyse 40 fecal samples collected clinically, and the detection results were consistent with cPCR. These results corroborate that the newly developed RPA method has good sensitivity and specificity and offers the advantage of short detection times, which can be harnessed for differential diagnosis and rapid detection of sp.
Topics: Humans; Animals; Recombinases; Blastocystis; Polymerase Chain Reaction; Nucleic Acid Amplification Techniques; Sensitivity and Specificity; Blastocystis Infections; Real-Time Polymerase Chain Reaction
PubMed: 37860882
DOI: 10.1017/S0031182023000975 -
The ISME Journal Dec 2023The soil amoeba Dictyostelium discoideum acts as both a predator and potential host for diverse bacteria. We tested fifteen Pseudomonas strains that were isolated from...
The soil amoeba Dictyostelium discoideum acts as both a predator and potential host for diverse bacteria. We tested fifteen Pseudomonas strains that were isolated from transiently infected wild D. discoideum for ability to escape predation and infect D. discoideum fruiting bodies. Three predation-resistant strains frequently caused extracellular infections of fruiting bodies but were not found within spores. Furthermore, infection by one of these species induces secondary infections and suppresses predation of otherwise edible bacteria. Another strain can persist inside of amoebae after being phagocytosed but is rarely taken up. We sequenced isolate genomes and discovered that predation-resistant isolates are not monophyletic. Many Pseudomonas isolates encode secretion systems and toxins known to improve resistance to phagocytosis in other species, as well as diverse secondary metabolite biosynthetic gene clusters that may contribute to predation resistance. However, the distribution of these genes alone cannot explain why some strains are edible and others are not. Each lineage may employ a unique mechanism for resistance.
Topics: Animals; Predatory Behavior; Dictyostelium; Pseudomonas; Amoeba; Bacteria
PubMed: 37884792
DOI: 10.1038/s41396-023-01535-5 -
PLoS Neglected Tropical Diseases Aug 2023In the human host, the protozoan parasite Entamoeba histolytica is adapted to a non-invasive lifestyle in the colon as well as to an invasive lifestyle in the...
In the human host, the protozoan parasite Entamoeba histolytica is adapted to a non-invasive lifestyle in the colon as well as to an invasive lifestyle in the mesenterial blood vessels and the liver. This means to cope with bacteria and human cells as well as various metabolic challenges. Galactose and N-acetylgalactosamine (GalNAc) are sugars of great importance for the amoebae, they attach to the host mucus and enterocytes via their well-studied Gal/GalNAc specific lectin, they carry galactose residues in their surface glycans, and they cleave GalNAc from host mucins. The enzyme UDP-glucose 4-epimerase (GalE) works as a bridge between the galactose and glucose worlds, it can help to generate glucose for glycolysis from phagocytosis products containing galactose as well as providing UDP-galactose necessary for the biosynthesis of galactose-containing surface components. E. histolytica contains a single galE gene. We recombinantly expressed the enzyme in Escherichia coli and used a spectrophotometric assay to determine its temperature and pH dependency (37°C, pH 8.5), its kinetics for UDP-glucose (Km = 31.82 μM, Vmax = 4.31 U/mg) and substrate spectrum. As observed via RP-HPLC, the enzyme acts on UDP-Glc/Gal as well as UDP-GlcNAc/GalNAc. Previously, Trypanosoma brucei GalE and the bloodstream form of the parasite were shown to be susceptible to the three compounds ebselen, a selenoorganic drug with antioxidant properties, diethylstilbestrol, a mimic of oestrogen with anti-inflammatory properties, and ethacrynic acid, a loop diuretic used to treat oedema. In this study, the three compounds had cytotoxic activity against E. histolytica, but only ebselen inhibited the recombinant GalE with an IC50 of 1.79 μM (UDP-Gal) and 1.2 μM (UDP-GalNAc), suggesting that the two other compounds are active against other targets in the parasite. The importance of the ability of GalE to interconvert UDP-GalNAc and UDP-GlcNAc may be that the trophozoites can generate precursors for their own cyst wall from the sugar subunits cleaved from host mucins. This finding advances our understanding of the biochemical interactions of E. histolytica in its colonic environment.
Topics: Humans; UDPglucose 4-Epimerase; Entamoeba histolytica; Galactose; Glucose; Acetylgalactosamine; Cysts; Uridine Diphosphate
PubMed: 37616327
DOI: 10.1371/journal.pntd.0011574 -
Veterinary Parasitology Jul 2024Blastocystis inhabits the digestive tracts of a diverse range of hosts. Transmission patterns, including host specificity, and the clinical and public health...
Blastocystis inhabits the digestive tracts of a diverse range of hosts. Transmission patterns, including host specificity, and the clinical and public health significance of Blastocystis in humans remain poorly understood. This study aimed to investigate the distribution and genetic diversity of Blastocystis in herbivorous and carnivorous reptiles in Eastern Thailand. A total of 501 faecal samples were collected from 363 iguanas, 79 bearded dragons, 50 tortoises, and nine snakes in an animal breeding farm in Chonburi Province, Eastern Thailand. Detection and differentiation of Blastocystis was based on amplification, sequencing, and phylogenetic analysis of specific small subunit (SSU) ribosomal RNA genes from faecal DNA extracted from the samples. Altogether 101/501 samples (20 %) were polymerase chain reaction (PCR) and sequencing-positive for Blastocystis, 90 (89 %) of which were from iguanas; the remaining positive samples were from African spurred tortoise (n=6), Bearded dragon (n=3), Leopard tortoise (n=1), and Red-footed tortoise (n=1). Phylogenetic analysis revealed that most of the Blastocystis sequences from iguanas were largely similar, and they were distinct from those of the tortoises. Subtype 17 was found in the three bearded dragons and likely reflected Blastocystis from prey animals. This is the largest survey of Blastocystis in reptiles to date. Remarkable differences in Blastocystis colonization rates and genetic diversity were observed between iguanas and other reptile orders, and what was considered Blastocystis colonization was only observed in herbivorous reptiles.
Topics: Animals; Blastocystis; Thailand; Genetic Variation; Blastocystis Infections; Host Specificity; Phylogeny; Feces; Reptiles; Turtles; Lizards; Snakes
PubMed: 38838380
DOI: 10.1016/j.vetpar.2024.110218 -
MBio Oct 2023Although most bacteria are quickly killed after phagocytosis by a eukaryotic cell, some pathogenic bacteria escape death after phagocytosis. Pathogenic species secrete...
Although most bacteria are quickly killed after phagocytosis by a eukaryotic cell, some pathogenic bacteria escape death after phagocytosis. Pathogenic species secrete polyP, and the polyP is necessary for the bacteria to prevent their killing after phagocytosis. Conversely, exogenous polyP prevents the killing of ingested bacteria that are normally killed after phagocytosis by human macrophages and the eukaryotic microbe . This suggests the possibility that in these cells, a signal transduction pathway is used to sense polyP and prevent killing of ingested bacteria. In this report, we identify key components of the polyP signal transduction pathway in . In cells lacking these components, polyP is unable to inhibit killing of ingested bacteria. The pathway components have orthologs in human cells, and an exciting possibility is that pharmacologically blocking this pathway in human macrophages would cause them to kill ingested pathogens such as .
Topics: Humans; Polyphosphates; Diphosphates; Dictyostelium; Bacteria; Phagocytosis; TOR Serine-Threonine Kinases
PubMed: 37754562
DOI: 10.1128/mbio.01939-23 -
Genes To Cells : Devoted To Molecular &... Dec 2023Cytokinesis, the final process of cell division, involves the accumulation of actin and myosin II filaments at the cell's equator, forming a contractile ring that...
Cytokinesis, the final process of cell division, involves the accumulation of actin and myosin II filaments at the cell's equator, forming a contractile ring that facilitates the division into two daughter cells. While light microscopy has provided valuable insights into the molecular mechanism of this process, it has limitations in examining individual filaments in vivo. In this study, we utilized transmission electron microscopy to observe actin and myosin II filaments in the contractile rings of dividing Dictyostelium cells. To synchronize cytokinesis, we developed a novel method that allowed us to visualize dividing cells undergoing cytokinesis with a frequency as high as 18%. This improvement enabled us to examine the lengths and alignments of individual filaments within the contractile rings. As the furrow constricted, the length of actin filaments gradually decreased. Moreover, both actin and myosin II filaments reoriented perpendicularly to the long axis during furrow constriction. Through experiments involving myosin II null cells, we discovered that myosin II plays a role in regulating both the lengths and alignments of actin filaments. Additionally, dynamin-like protein A was found to contribute to regulating the length of actin filaments, while cortexillins were involved in regulating their alignment.
Topics: Actomyosin; Actins; Dictyostelium; Actin Cytoskeleton; Cytokinesis; Myosin Type II
PubMed: 37844904
DOI: 10.1111/gtc.13073 -
Mikrobiyoloji Bulteni Apr 2024Blastocystis spp. are the most common intestinal protozoan parasites detected in human stool samples. While identified long before today, its pathogenicity remains...
Blastocystis spp. are the most common intestinal protozoan parasites detected in human stool samples. While identified long before today, its pathogenicity remains controversial. It is generally asymptomatic but in symptomatic cases, many gastrointestinal symptoms, especially diarrhea, have been associated with Blastocystis infection. In recent years, the relationship between the symptoms observed in cases and Blastocystis subtypes (ST) has been reported. The aim of this study was to detect Blastocystis in diarrheal cases admitted to the Aydın Adnan Menderes University Faculty of Medicine, Department of Parasitology Laboratory, to determine subtypes and allele diversity and to investigate its relationship with clinical symptoms. For this purpose, diarrheal stool samples of 200 cases were included in the study and their demographic characteristics (age, gender, residence) and clinical findings (abdominal pain, dyspepsia, nausea-vomiting, weakness, weight loss, anal itching, rash, urticaria) were recorded. Blastocystis was detected by direct microscope method (DM) and by molecular analyses which were performed with polymerase chain reaction (PCR). Subtype diversity was determined based on DNA sequence analysis by PCR targeting the Blastocystis ribosomal ribonucleic acid small subunit (SSU rRNA) gene. In addition, alleles related to Blastocystis subtypes were determined and statistically compared between all data and clinical findings. In the current study, Blastocystis was detected in 31 (15.5%) samples by DM and in 35 (17.5%) samples by PCR specific to the Blastocystis SSU rRNA gene among 200 diarrheal stool samples. No statistical difference was detected between Blastocystis and demographic characteristics. Dyspepsia and nausea-vomiting symptoms differed significantly in cases with Blastocystis compared to negative ones (p= 0.0025, p= 0.0498). Blastocystis subtype was detected in 33 samples by SSU rRNA sequence analysis, and the subtype distribution was ST1 (n= 10, 30.3%), ST2 (n= 4, 12.1%) and ST3 (n= 19, 57.6%). In the statistical evaluation between clinical findings and Blastocystis subtypes, a relationship was found between dyspepsia and Blastocystis ST3 (p= 0.0039). The allele diversity of Blastocystis subtypes was determined as allele 4 (10/10) in all ST1, allele 11 (2/4) and 12 (2/4) in ST2, allele 34 (14/19), 36 (4/19), and 38 (1/19) in ST3. In conclusion, our study provides important data on the molecular epidemiological characteristics of the Blastocystis by determining positivity, subtypes and alleles in diarrheal cases. Therefore, within the scope of the one health approach, comprehensive molecular epidemiological studies are required to determine the presence and genotypes of Blastocystis in human, animal and environmental samples.
Topics: Humans; Blastocystis; Blastocystis Infections; Diarrhea; Male; Genetic Variation; Female; Adult; Feces; Alleles; Middle Aged; Adolescent; Young Adult; Child; Aged; Child, Preschool; Polymerase Chain Reaction; DNA, Protozoan; Turkey
PubMed: 38676586
DOI: 10.5578/mb.202498207 -
Scientific Reports May 2024The patterns of Formin B and of the Arp2/3 complex formed during mitosis were studied in a mutant of Dictyostelium discoideum that produces multinucleate cells, which...
The patterns of Formin B and of the Arp2/3 complex formed during mitosis were studied in a mutant of Dictyostelium discoideum that produces multinucleate cells, which divide by the ingression of unilateral cleavage furrows. During cytokinesis the cells of this mutant remain spread on a glass surface where they generate a planar pattern based on the sorting-out of actin-binding proteins. During anaphase, Formin B and Arp2/3 became localized to the regions of microtubule asters around the centrosomes; Formin B in particular in the form of round, quite uniformly covered areas. These areas have been shown to be depleted of myosin II and the actin-filament crosslinker cortexillin, and to be avoided by cleavage furrows on their path into the cell.
Topics: Microtubules; Dictyostelium; Mitosis; Microfilament Proteins; Actin-Related Protein 2-3 Complex; Protozoan Proteins; Protein Transport; Cytokinesis; Actins
PubMed: 38755233
DOI: 10.1038/s41598-024-61967-7