Background: It has been known for quite some time now that silver nanoparticles (AgNP) can inhibit microbial growth and even kill microbes. Our investigation reports the antimicrobial activity of AgNP against a model bacterium, Escherichia coli.
Entamoeba histolytica, the causative agent of amoebiasis, does not form cysts in vitro, so reptilian pathogen Entamoeba invadens is used as an Entamoeba encystation model. During the in vitro encystation of E. invadens, a few multinucleated giant cells (MGC) were also appeared in the culture along with cysts. Like the cyst, these MGC's were also formed in the multicellular aggregates found in the encystation culture. Time-lapse live cell imaging revealed that MGC's were the result of repeated cellular fusion with fusion-competent trophozoites as a starting point. The early MGC were non-adherent, and they moved slowly and randomly in the media, but under confinement, MGC became highly motile and directionally persistent. The increased motility resulted in rapid cytoplasmic fissions, which indicated the possibility of continuous cell fusion and division taking place inside the compact multicellular aggregates. Following cell fusion, each nucleus obtained from the fusion-competent trophozoites gave rise to four nuclei with half genomic content. All the haploid nuclei in MGC later aggregated and fused to form a polyploid nucleus. These observations have important implications on Entamoeba biology as they point toward the possibility of E. invadens undergoing sexual or parasexual reproduction.
Entamoeba histolytica and its reptilian counterpart and encystation model Entamoeba invadens formed a polarized monopodial morphology when treated with pentoxifylline. This morphology was propelled by retrograde flow of the cell surface resulting from a cyclic sol-gel conversion of cytoplasm and a stable bleb at the leading edge. Pentoxifylline treatment switched the unpolarized, adherent trophozoites to the nonadherent, stable bleb-driven form and altered the motility pattern from slow and random to fast, directionally persistent, and highly chemotactic. Interestingly, exogenously added adenosine produced multiple protrusions and random motility, an opposite phenotype to that of pentoxifylline. Thus, pentoxifylline, an adenosine antagonist, may be inducing the monopodial morphology by preventing lateral protrusions and restricting the leading edge to one site. The polarized form of E. invadens was aggregation competent, and time-lapse microscopy of encystation revealed its appearance during early hours, mediating the cell aggregation by directional cell migration. The addition of purine nucleotides to in vitro encystation culture prevented the formation of polarized morphology and inhibited the cell aggregation and, thus, the encystation, which further showed the importance of the polarized form in the Entamoeba life cycle. Cell polarity and motility are essential in the pathogenesis of Entamoeba parasites, and the stable bleb-driven polarized morphology of Entamoeba may also be important in invasive amoebiasis.
21The cyst wall of Entamoeba histolytica, the causative agent of the amoebiasis, is a 22 potential target for new drugs. The "Wattle and Daub" model of cyst wall formation of 23 Entamoeba invadens had already been reported. In this study, we demonstrate in more detail the 24 morphological stages of chitin wall formation in E. invadens using fluorescent chitin-binding 25 dyes and immunolocalization of the cyst wall proteins. Here, the expression and localization of 26 chitin synthase and the importance of actin cytoskeleton dynamics at cellular level, during 27 encystations have been demonstrated for the first time. Chitin deposition was found to be 28 initiated on the cell surface mostly from one distinct point, though multipoint initiation was also 29 observed sometimes. From these points, the wall grew outwards and gradually covered the entire 30 cyst surface with time. The initiation of chitin deposition was guided by the localization of chitin 31 synthase 1 on the plasma membrane. The gradual formation of the cyst wall follows the Wattle 32 and daub model. The chitin deposition occurred on the foundation of Jacob lectin at the cell 33 membrane, and the other cyst wall components, like chitinase, and Jessie were also found to be 34 present in the growing incomplete walls. In contrary to the Wattle and daub model, Jessie was 35 found to be expressed and localized in the growing wall at the early hours of encystations. 36During encystation, F-actin was reorganized into the cortical region within an hour encystation 37 initiation and remained intact until the completion of the chitin wall. Disruption of cortical actin 38 polymerization with 2, 3-Butanedione monoxime inhibited proper wall formation but produced 39 wall-less cysts or cysts with defective chitin wall. Malformations of cyst-walls were mainly due 40 to improper localization and activity of chitin synthases, which indicates the indispensability of 41 cortical actin cytoskeleton for the proper cyst wall formation.42 3 43 Author Summary 44 Entamoeba parasites reach new hosts using the resistant cyst form, so preventing its 45 formation can stop the spread of amoebiasis. The resistant nature of the cyst is due to the chitin 46 wall, and thus identifying the critical steps of wall formation could provide targets for designing 47 new drugs. Here we studied the morphological stages of the cyst wall formation by observing 48 how the chitin and other cell wall components were deposited on the cell surface using 49 fluorescent chitin-binding dyes and antibodies against cyst wall proteins. In most cases, the 50 chitin wall was found to start from one distinct point from which it spread all over the cell 51 surface, guided by chitin synthase. The composition of these incomplete walls was the same as a 52 mature cyst wall indicating that the wall may be a result of extracellular self-assembly of its 53 constituents from one starting point. We have also observed that F-actin polymerized in the 54 cortex of encysting cells and its disruption resulted in wall-less...
The inability of enteric pathogens to produce butyrate may impair epithelial cell function, whereas production of D-lactate may enhance mucosal damage in diarrhoeal disease. The presence of luminal starch may be helpful in shifting the fermentation profile to a more favourable pattern.
statementPentoxifylline, the adenosine receptor antagonist induced a stable bleb driven polarized morphology in Entamoeba characterized by fast, directionally persistent and highly chemotactic motility. AbstractProtozoan parasites Entamoeba histolytica and Entamoeba invadens formed a polarized phenotype, an elongated shape with a single leading edge and a trailing edge when treated with pentoxifylline. The leading edge of the polarized morphology was a spherical protrusion devoid of F-actin but with occasional F-actin scars, indicating the presence of bleb. The polarized form was stable bleb driven since the blebbing was limited to the leading edge. Pentoxifylline induced chemokinesis in Entamoeba as it switched the motility pattern from slow and random to fast and directionally persistent. Pentoxifylline speeded up the cell aggregation in E. invadens during growth and encystation due to enhanced chemotaxis of the polarized form. The transformation of non-polarized adherent trophozoites to nonadherent stable bleb driven form occurred via lamellipodial and bleb driven adherent intermediate phenotypes. The nonadherent polarized phenotype was highly motile under confinement and moved by rearward plasma membrane flow.In contrast to pentoxifylline, adenosine, the adenosine receptor agonist, stimulated the formation of multiple protrusions leading to random motility. Thus pentoxifylline might prevent lateral protrusions by inhibiting adenosine receptor, producing the monopodial polarized morphology.Cell migration is an important event in embryonic development, wound healing, immune response, and cancer progression. The first step in cell migration is the polarization during which the cells break symmetry to form a leading edge and a trailing edge. Depending on the leading edge, motility can be of two types, lamellipodial motility driven by actin polymerization and pressure driven blebbing. Cells like keratocytes and fibroblasts use lamellipodia at the leading edge while primordial germ cells and amoeboid tumor cells use blebbing motility (Blaser et al., 2006;Paňková et al., 2010). Bleb driven cell migration have been reported in many biological events like embryonic development, Dictyostelium chemotaxis, and cancer metastasis (Fackler and Grosse, 2008) but unlike lamellipodial motility, its mechanics is not well understood.Enteric parasite Entamoeba histolytica is a highly motile organism, and it has been reported to use only the bleb driven motility both in vitro and in vivo and thus can be considered as an important model to study blebbing (Maugis et al., 2010). As shown previously in progenitor cells (Diz-Muñoz et al., 2010), blebbing reduced directional persistence in E.histolytica. Here we report that both human pathogen E. histolytica and its reptilian counterpart and encystation model Entamoeba invadens readily formed a fast moving, elongated morphology when treated with a millimolar concentration of methylxanthines like pentoxifylline or caffeine.This morphology showed a single leading edge with stable bleb, ma...
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