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The intestinal parasites Giardia lamblia and Entamoeba histolytica are major causes of morbidity and mortality associated with diarrheal diseases. Metronidazole is the most common drug used to treat giardiasis and amebiasis. Despite its efficacy, treatment failures in giardiasis occur in up to 5%-40% of cases. Potential resistance of E. histolytica to metronidazole is an increasing concern. Therefore, it is critical to search for more effective drugs to treat giardiasis and amebiasis. We identified antigiardial and antiamebic activities of the rediscovered nitroimidazole compound, fexinidazole, and its sulfone and sulfoxide metabolites. Fexinidazole is equally active against E. histolytica and G. lamblia trophozoites, and both metabolites were 3- to 18-fold more active than the parent drug. Fexinidazole and its metabolites were also active against a metronidazole-resistant strain of G. lamblia. G. lamblia and E. histolytica cell extracts exhibited decreased residual nitroreductase activity when metabolites were used as substrates, indicating nitroreductase may be central to the mechanism of action of fexinidazole. In a cell invasion model, fexinidazole and its metabolites significantly reduced the invasiveness of E. histolytica trophozoites through basement membrane matrix. A q.d. oral dose of fexinidazole and its metabolites at 10 mg/kg for 3 days reduced G. lamblia infection significantly in mice compared to control. The newly discovered antigiardial and antiamebic activities of fexinidazole, combined with its FDA-approval and inclusion in the WHO Model List of Essential Medicines for the treatment of human African trypanosomiasis, offer decreased risk and a shortened development timeline toward clinical use of fexinidazole for treatment of giardiasis or amebiasis.

The anaphase promoting complex (APC or cyclosome) is a major ubiquitin ligase that coordinates mitotic and G1 progression, acting as a major regulator of chromosome segregation. While the human APC contains fourteen subunits, it is yet to be explored in the pathogen Entamoeba histolytica. Our study reveals the existence of a single functional Apc10 homolog in E. histolytica, which acts as a processivity factor of ubiquitin ligase activity in human. A cDNA library generated from HM1:IMSS strain of E. histolytica was screened for interaction partners of EhApc10 in yeast two hybrid study. The novel interactor, a glycolytic enzyme, pyruvate phosphate dikinase (Ppdk) was found to interact with EhApc10 and further validated by in vitro assay. A comprehensive in silico study has emphasized the structural and functional aspects, encompassing physicochemical traits, predictive 3D structure modelling, validation of EhApc10-EhPpdk interaction through molecular docking and simulation. The interplay between a cell cycle protein and a glycolytic enzyme highlights the connection between cellular metabolism and the cell cycle regulatory mechanism. The study serves as the groundwork for future research on the non-mitotic role of APC beyond cell cycle. [ABSTRACT FROM AUTHOR]

Copyright of Biomédica: Revista del Instituto Nacional de Salud is the property of Instituto Nacional de Salud of Colombia and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Entamoeba histolytica, a protozoan parasite, is the causative agent of amoebiasis, which is a significant global health concern. The virulence mechanisms underlying its pathogenicity are multifaceted and complex. However, endocytic processes and motility are well accepted virulence determinants. As previously reported, an AGCK family kinase, EhAGCK1 to be involved in trogocytosis exclusively while another one from same family named EhAGCK2 participates in all actin dependent endocytic processes. As the kinase dead mutants of EhAGCK1 showed significant defect in destruction of live host cells and also the localisation pattern of same is distinguishable from EhAGCK2. From observations so far, it appears that former initiates a distinguishable signaling cascade. In this work, we have demonstrated distinct biochemical properties of kinases involved in related yet distinguishable endocytic processes for the first time. Our biochemical characterization highlights distinct ion dependency of EhAGCK1 along with substrate specificity. We also show upstream activator of these kinases, 3-phosphoinositide dependent kinase 1 (PDK1) activity and its role in activating the kinase activity. The kinases exhibit property of autophosphorylation, and which may regulate the kinase activity subsequently. Summarily, these studies show that EhAGCK1 and EhAGCK2 show distinct biochemical properties which further confirm their unique role in related endocytic processes of trogocytosis and phagocytosis. Author summary: The protozoan parasite E. histolytica causes amebiasis in humans. The trophozoites depend on variety of endocytic processes to survive and cause pathogenesis in the gut of host system. Phagocytosis and trogocytosis are two related endocytic processes known to contribute to the pathogenesis by the parasite in vivo. The two processes are phenotypically different, trogocytosis being fast paced resembling nibbling and specific to live host cells. In terms of molecular mechanism of these processes, an AGC family kinase, EhAGCK1 is known to be involved in trogocytosis specifically while another one EhAGCK2 is involved in all types of endocytic processes including fluid uptake. Our biochemical characterization of these two kinases reveal that they have distinct substrate specificity and ion requirement. The overall biochemical assessment indicates the two kinases belonging to same family are involved in the phagocytic and trogocytic pathway distinctly. Hence, although the phagocytosis and trogocytosis at molecular level appear to be related but they also involve steps unique to each pathway. [ABSTRACT FROM AUTHOR]

Although diloxanide is a drug of choice for treating asymptomatic amoebiasis, its mechanism of action (MOA) remains unclear. This review aims to shed light on the current understanding of the effectiveness and MOA of diloxanide in treating amoebiasis . It involves analysis of articles, retrieved from PubMed, Google Scholar and EBSCOhost, on diloxanide and the treatment of Entamoeba histolytica infection. Diloxanide is used in an ester form, which allows its high luminal concentration and greater efficacy than metronidazole in the management of asymptomatic amoebiasis. The current understanding of the action of diloxanide is based on its structural similarity to chloramphenicol at dichloroacetamide group. It acts against protein synthesis in E. histolytica trophozoites, blocking their conversion to more virulent and invasive cyst forms. Furthermore, it has a parasite clearance rate of 81–96% and treats amoebic abscesses when combined with metronidazole and chloroquine. Nevertheless, it is associated with adverse events such as flatulence, anorexia, headache and urticaria. Diloxanide is efficacious against amoebiasis but there is a need to explore its structure–activity relationship.The study suggests future directions, including novel drug formulations, diagnostic improvements, and combination regimens to enhance treatment outcomes and mitigate relapse associated with the use of diloxanide. [ABSTRACT FROM AUTHOR]

Diabetic kidney disease (DKD) is the main cause of deaths due to diabetes mellitus (DM). Due to the complexity of its onset, it is difficult to achieve accurate prevention and treatment. The classically activated macrophage (M1) polarization is a crucial proinflammatory mechanism of DKD, while the interaction and cascade effects of oxidative stress and inflammatory response remain to be elucidated. A urine proteomic analysis of patients with DM indicated that peroxiredoxin 2 (PRDX2) had the higher abundance in DKD. We recently found that PRDX of parasitic protozoa Entamoeba histolytica, which was similar to human PRDX2 in amino acid sequence and spatial structure, could activate the inflammatory response of macrophages through toll-like receptor 4 (TLR4). Hence, our study was designed to explore the role of PRDX2 in chronic inflammation during DKD. Combined with in vivo and in vitro experiments, results showed that the PRDX2 was positively correlated with DKD progression and upregulated by high glucose or recombinant tumor necrosis factor-α in renal tubular epithelial cells; Besides, recombinant PRDX2 could promote M1 polarization of macrophages, and enhance the migration as well as phagocytic ability of macrophages through TLR4. In summary, our study has explored the novel role of PRDX2 in DKD to provide a basis for further research on the diagnosis and treatment of DKD. [ABSTRACT FROM AUTHOR]

Entamoeba histolytica causes amebiasis, a significant global health issue, with millions affected annually, especially in developing countries. EhDUF2419, an important protein involved in E. histolytica's queuine salvage pathway and its interaction network, remains unclear. To explore this, we transfected E. histolytica trophozoites with a plasmid encoding Myc-tagged EhDUF2419 and achieved successful overexpression. Through immunoprecipitation with the Myc antibody followed by mass spectrometry, we identified 335 proteins interacting with Myc-tagged EhDUF2419, including over 100 ribosomal proteins, along with translation initiation and elongation factors, and aminoacyl-tRNA synthetases. Ribosome purification revealed the presence of EhDUF2419 in ribosomal protein-enriched fractions. Treatment with queuosine (Q) significantly reduced the EhDUF2419 protein levels and decreased the Q-modified tRNA in Myc-tagged EhDUF2419 overexpressing trophozoites. This effect, which was Q-dependent, was not observed in strains carrying an empty vector control or overexpressing a truncated form of EhDUF2419 lacking catalytic activity. The reduction in the EhDUF2419 protein levels was regulated by proteasome-mediated degradation, as evidenced by the reduced degradation in the presence of MG132, a proteasome inhibitor. Our study uncovers the novel interaction of EhDUF2419 with ribosomal proteins and its regulation by the proteasome machinery, providing new insights into its role in E. histolytica and potential therapeutic strategies. [ABSTRACT FROM AUTHOR]

Macropinocytosis is an evolutionarily conserved endocytic process that plays a vital role in internalizing extracellular fluids and particles in cells. This non-selective endocytic pathway is crucial for various physiological functions such as nutrient uptake, sensing, signaling, antigen presentation, and cell migration. While macropinocytosis has been extensively studied in macrophages and cancer cells, the molecular mechanisms of macropinocytosis in pathogens are less understood. It has been known that Entamoeba histolytica, the causative agent of amebiasis, exploits macropinocytosis for survival and pathogenesis. Since macropinocytosis is initiated by actin polymerization, leading to the formation of membrane ruffles and the subsequent trapping of solutes in macropinosomes, actin cytoskeleton regulation is crucial. Thus, this study focuses on unraveling the role of well-conserved actin cytoskeleton regulators, Rho small GTPase family proteins, in macropinocytosis in E. histolytica. Through gene silencing of highly transcribed Ehrho/Ehrac genes and following flow cytometry analysis, we identified that silencing EhracM enhances dextran macropinocytosis and affects cellular migration persistence. Live imaging and interactome analysis unveiled the cytosolic and vesicular localization of EhRacM, along with its interaction with signaling and membrane traffic-related proteins, shedding light on EhRacM's multiple roles. Our findings provide insights into the specific regulatory mechanisms of macropinocytosis among endocytic pathways in E. histolytica, highlighting the significance of EhRacM in both macropinocytosis and cellular migration. Author summary: Entamoeba histolytica is an intestinal protozoan parasite that causes amoebic dysentery and liver abscesses in humans. This organism exploits macropinocytosis, a cellular process that engulfs extracellular fluids and particles, for its survival and pathogenicity. Although macropinocytosis is well-characterized in immune cells and cancer cells as it is essential for nutrient uptake, its mechanisms in pathogens, such as E. histolytica, remain less explored. Our research focused on the molecular mechanisms underpinning macropinocytosis in this parasite, specifically examining the role of Rho small GTPase family proteins. These proteins are critical regulators of the actin cytoskeleton in eukaryotic cells. Our study reveals that one specific Rho small GTPase, EhRacM, is in the maturation of macropinosomes as well as in directing linear cell migration. The physiological significance of EhRacM in regulating both macropinocytosis and migration opens new avenues for understanding the role of Rho small GTPases in these signaling pathways, which could eventually lead to the development of new control measures against diseases caused by this parasite. [ABSTRACT FROM AUTHOR]

Entamoeba nuttalli is genetically the closest to Entamoeba histolytica, the causative agent of human amebiasis. E. nuttalli is found in Macaca species, exhibiting no symptoms while potentially virulent. Using comparative genomics of Entamoeba species, we identified a gene encoding an E. nuttalli‐specific protein containing 42 repeats of an octapeptide (PTORS). In the present study, we analyzed the genes in E. nuttalli strains derived from various geographic locations and host species. Sequence analysis of genomic DNA from four strains indicated 43, 44, and 48 repeat types in addition to 42 repeats and remarkable genetic diversity in the repeat region, although all nucleotide substitutions were synonymous. In contrast, the sequences of the N‐terminal side region and C‐terminus were identical among the strains. Monoclonal antibodies prepared against recombinant PTORS were reactive to the repeat regions but not to the N‐terminal side regions. Polyclonal antibodies did not react with the N‐terminal region, demonstrating that the repeat region had higher antigenicity. Analysis using synthetic peptides revealed that the two repeats of the octapeptide functioned as epitopes. Immunofluorescence microscopy using monoclonal antibodies demonstrated the surface localization of PTORS. These results suggest that the repeat region of PTORS plays an important role in host–parasite interactions. [ABSTRACT FROM AUTHOR]

Entamoeba histolytica causes invasive amoebiasis, an important neglected tropical disease with a significant global health impact. The pathogenicity and survival of E. histolytica and its reptilian equivalent, Entamoeba invadens, relies on its ability to exhibit efficient motility, evade host immune responses, and exploit host resources, all of which are governed by the actin cytoskeleton remodeling. Our study demonstrates the early origin and the regulatory role of TALE homeobox protein EiHbox1 in actin‐related cellular processes. Several genes involved in different biological pathways, including actin dynamics are differentially expressed in EiHbox1 silenced cells. EiHbox1 silenced parasites showed disrupted F‐actin organization and loss of cellular polarity. EiHbox1's presence in the anterior region of migrating cells further suggests its involvement in maintaining cellular polarity. Loss of polarized morphology of EiHbox1 silenced parasites leads to altered motility from fast, directionally persistent, and highly chemotactic to slow, random, and less chemotactic, which subsequently leads to defective aggregation during encystation. EiHbox1 knockdown also resulted in a significant reduction in phagocytic capacity and poor capping response. These findings highlight the importance of EiHbox1 of E. invadens in governing cellular processes crucial for their survival, pathogenicity, and evasion of the host immune system. [ABSTRACT FROM AUTHOR]

Recent studies have shown that extracellular vesicles (EVs) secreted by various parasites are capable of modulating the host's innate immune responses, such as by altering macrophage (Mϕ) phenotypes and functions. Studies have shown that Mϕ promote early host responses to amoebic infection by releasing proinflammatory cytokines that are crucial to combating amoebiasis. Here, we are reporting for the first time the effect of EVs released by Entamoeba histolytica (EhEVs) on human THP‐1 differentiated Mϕ (THP‐1 Mϕ). We show that the EhEVs are internalized by THP‐1 Mϕ which leads to differential regulation of various cytokines associated with both M1 and M2 Mϕ. We also saw that EhEV treatment thwarted Type 2 immune‐response‐related transcriptome pSTAT6 in the THP‐1 Mϕ. Furthermore, EhEVs stimulated Mϕ to reduce their energy demand by suppressing oxidative phosphorylation (OXPHOS) and adenosine triphosphate (ATP) production. Hence, the human parasite E. histolytica–derived EVs are capable of eliciting an immune response from Mϕ that may contribute to overall infection status. [ABSTRACT FROM AUTHOR]

Intestinal parasites are part of the intestinal ecosystem and have been shown to establish close interactions with the intestinal microbiota. However, little is known about the influence of intestinal protozoa on the regulation of the immune response. In this study, we analyzed the regulation of the immune response of germ-free mice transplanted with fecal microbiota (FMT) from individuals with multiple parasitic protozoans (P) and non-parasitized individuals (NP). We determined the production of intestinal cytokines, the lymphocyte populations in both the colon and the spleen, and the genetic expression of markers of intestinal epithelial integrity. We observed a general downregulation of the intestinal immune response in mice receiving FMT-P. We found significantly lower intestinal production of the cytokines IL-6, TNF, IFN-γ, MCP-1, IL-10, and IL-12 in the FMT-P. Furthermore, a significant decrease in the proportion of CD3+, CD4+, and Foxp3+ T regulatory cells (Treg) was observed in both, the colon and spleen with FMT-P in contrast to FMT-NP. We also found that in FMT-P mice there was a significant decrease in tjp1 expression in all three regions of the small intestine; ocln in the ileum; reg3γ in the duodenum and relmβ in both the duodenum and ileum. We also found an increase in colonic mucus layer thickness in mice colonized with FMT-P in contrast with FMT-NP. Finally, our results suggest that gut protozoa, such as Blastocystis hominis, Entamoeba coli, Endolimax nana, Entamoeba histolytica/E. dispar, Iodamoeba bütschlii, and Chilomastix mesnili consortia affect the immunoinflammatory state and induce functional changes in the intestine via the gut microbiota. Likewise, it allows us to establish an FMT model in germ-free mice as a viable alternative to explore the effects that exposure to intestinal parasites could have on the immune response in humans. [ABSTRACT FROM AUTHOR]

In humans and Drosophila melanogaster, the functional convergence of the endosomal sorting complex required for transport (ESCRT) machinery that is in charge of selecting ubiquitinated proteins for sorting into multivesicular bodies, and the retromer, that is the complex responsible for protein recycling to the plasma membrane and Golgi apparatus. ESCRT and retromer complexes are codependent for protein sorting recycling, degradation, and secretion. In this article, we studied the EhVps35 C isoform (referred to as EhVps35), that is the central member of the Entamoeba histolytica retromer, and its relation with the ESCRT machinery during sorting and protein recycling events and their involvement virulence. Our findings revealed that EhVps35 interacts with at least 300 proteins that participate in multiple cellular processes. Laser confocal and transmission electronic microscopy images, as well as secretion assays, revealed that EhVps35 is secreted in vesicles together with EhVps23 and EhADH (both ESCRT machinery proteins). In addition, immunoprecipitation, immunofluorescence, and molecular docking assays revealed the relationship among EhVps35 and other ESCRT machinery proteins. Red blood cell stimulus increased EhVps35 secretion, and the knockdown of the Ehvps35 gene in trophozoites reduced their capacity to migrate and invade tissues. This also impacts the cellular localization of ubiquitin, EhVps23 (ESCRT-I), and EhVps32 (ESCRT-III) proteins, strongly suggesting their functional relationship. Our results, taken together, give evidence that EhVps35 is a key factor in E. histolytica virulence mechanisms and that it, together with the ESCRT machinery components and other regulatory proteins, is involved in vesicle trafficking, secretion, migration, and cell proliferation. [ABSTRACT FROM AUTHOR]

Several studies with kaempferol (KP) and linearolactone (LL) have demonstrated their antiparasitic activity. However, the toxicity of these treatments is unknown. Therefore, this study aimed to evaluate the possible toxicological effects of intraperitoneal (i.p.) administration of KP or LL on the amoebic liver abscess model (ALA) in Mesocricetus auratus. An ALA was induced in male hamsters with 1.5 × 105Entamoeba histolytica (E. histolytica) trophozoites inoculated in the left hepatic lobe. The lesion evolved for 4 days, and then KP (5 mg/kg body weight/day) or LL (10 mg/kg body weight/day) was administered for 4 consecutive days. Then, magnetic resonance imaging (MRI), paraclinical analyses, and necropsy for histopathological evaluation were performed. There was similar ALA inhibition by KP (19.42%), LL (28.16%), and metronidazole, the antiamoebic control (20.87%) (p ≤ 0.05, analysis of variance [ANOVA]). There were hepatic and renal biochemical alterations in all treatment groups, mainly for KP (aspartate aminotransferase: 347.5 ± 37.5 U/L; blood urea nitrogen: 19.4 ± 1.9 g/dL; p ≤ 0.05, ANOVA). Lesions found in the organs were directly linked to the pathology. In conclusion, KP and LL decreased ALA development and exerted fewer toxicological effects compared with metronidazole. Therefore, both compounds exhibit therapeutic potential as an alternative treatment of amoebiasis caused by E. histolytica. However, additional clinical studies in different contexts are required to reaffirm this assertion. [ABSTRACT FROM AUTHOR]

Tetraspanins (TSPANs) are a family of proteins highly conserved in all eukaryotes. Although protein-protein interactions of TSPANs have been well established in eukaryotes including parasitic protists, the role they play in parasitism and pathogenesis remains largely unknown. In this study, we characterized three representative members of TSPANs, TSPAN4, TSPAN12, and TSPAN13 from the human intestinal protozoan Entamoeba histolytica. Co-immunoprecipitation assays demonstrated that TSPAN4, TSPAN12 and TSPAN13 are reciprocally pulled down together with several other TSPAN-interacting proteins including TSPAN binding protein of 55kDa (TBP55) and interaptin. Blue native-PAGE analysis showed that these TSPANs form several complexes of 120–250 kDa. Repression of tspan12 and tspan13 gene expression led to decreased secretion of cysteine proteases, while repression of tspan4 led to a four-fold increase in the activity of cysteine proteases in crude extracellular vesicles (EVs) fraction. Meanwhile, strains overexpressing HA-tagged TSPAN12 and TSPAN13 demonstrated reduced adhesion to collagen. Altogether, this study reveals that the TSPANs, especially TSPAN12 and TSPAN13, are engaged with complex protein-protein interactions and are involved in the pathogenicity-related biological functions such as protease secretion and adhesion, offering insights into the potential regulatory mechanisms of tetraspanins in protozoan parasites. Author summary: Tetraspanins (TSPANs) are involved in critical roles such as cellular communication, adhesion, and immune responses. However, their function in parasitic protozoa, particularly in relation to disease-causing mechanisms, remains poorly understood. Our study explores the role of three tetraspanins (TSPAN4, TSPAN12, and TSPAN13) in Entamoeba histolytica, an intestinal parasite which causes amebiasis, and a significant health problem affecting millions of people worldwide. Our findings reveal that these amebic TSPANs form various protein complexes similar to what is observed in other organisms. We also found that amebic TSPANs influence crucial processes related to the parasite's pathogenicity, such as adhesion to collagen and secretion of cysteine proteases (CPs). Gene silencing of TSPAN12 and TSPAN13 caused reduction in CP secretion, while overexpression resulted in decreased adhesion. Additionally, TSPAN4 appears to play a distinct role in exosome-mediated pathways. Our findings provide insights into a potential alternative mechanism of CP secretion via exosomes, in which amebic TSPANs may play a role. [ABSTRACT FROM AUTHOR]

Vegetables, especially those eaten raw, have been implicated in several foodborne disease outbreaks. Since multiple vegetable matrices and hazards are involved, risk managers have to prioritize those with the greatest impact on public health to design control strategies. In this study, a scientific-based risk ranking of foodborne pathogens transmitted by leafy green vegetables in Argentina was performed. The prioritization process included hazard identification, evaluation criteria identification and definition, criteria weighting, expert survey design and selection and call for experts, hazard score calculation, hazard ranking and variation coefficient, and result analysis. Regression tree analysis determined four risk clusters: high (Cryptosporidum spp., Toxoplasma gondii, Norovirus), moderate (Giardia spp., Listeria spp., Shigella sonnei), low (Shiga toxin-producing Escherichia coli, Ascaris spp., Entamoeba histolytica, Salmonella spp., Rotavirus, Enterovirus) and very low (Campylobacter jejuni, hepatitis A virus and Yersinia pseudotuberculosis). Diseases caused by Norovirus, Cryptosporidium spp. and T. gondii do not require mandatory notification. Neither viruses nor parasites are included as microbiological criteria for foodstuff. The lack of outbreak studies did not allow to accurately identify vegetables as a source of Norovirus disease. Information on listeriosis cases or outbreaks due to vegetable consumption was not available. Shigella spp. was the main responsible for bacterial diarrhea, but it has not been epidemiologically associated with vegetable consumption. The quality of the available information for all hazards studied was very low and low. The implementation of good practice guidelines throughout the entire vegetable production chain could prevent the presence of the identified hazards. The current study allowed the identification of vacancy areas and could help reinforce the need for performing epidemiological studies on foodborne diseases potentially associated with vegetable consumption in Argentina. [ABSTRACT FROM AUTHOR]

Increasing awareness regarding health promotion and disease prevention has driven inclusion of fermented foods and beverages in the daily diet. These are the enormous sources of beneficial microbes, probiotics. This study aims to isolate yeast strains having probiotic potential and effectivity against colitis. Initially, ninety-two yeast strains were isolated from Haria, an ethnic fermented beverage of West Bengal, India. Primary screening was done by their acid (pH 4) and bile salt (0.3%) tolerance ability. Four potent isolates were selected and found effective against Entamoeba histolytica, as this human pathogen is responsible to cause colitis. They were identified as Saccharomyces cerevisiae. They showed luxurious growth even at 37 oC, tolerance up to 5% of NaCl, resistance to gastric juice and high bile salt (2.0%) and oro-gastrointestinal transit tolerance. They exhibited good auto-aggregation and co-aggregation ability and strong hydrophobicity. Finally, heat map and principal component analysis revealed that strain Y-89 was the best candidate. It was further characterised and found to have significant protective effects against DSS-induced colitis in experimental mice model. It includes improvement in colon length, body weight and organ indices; reduction in disease activity index; reduction in cholesterol, LDL, SGPT, SGOT, urea and creatinine levels; improvement in HDL, ALP, total protein and albumin levels; decrease in coliform count and restoration of tissue damage. This study demonstrates that the S. cerevisiae strain Y-89 possesses remarkable probiotic traits and can be used as a potential bio-therapeutic candidate for the prevention of colitis. [ABSTRACT FROM AUTHOR]

Background: Entamoeba histolytica is a parasitic organism that infects the intestines and causes dysentery, characterized by intestine inflammation. Objectives: We aimed to estimate calprotectin levels and their relationship to the severity and progression of intestinal inflammation. Methods: We conducted a cross-sectional study in Al-Habbobi Teaching Hospital Thi-Qar, Al-Nasseriah, Iraq, from March 10 to October 1, 2023. The sample included 50 men and 50 women aged in three groups: 25-29, 30-34, and >34 years. We employed microscopy, direct wet method, and formal-ether concentration approach to detect parasites. Also, we employed nzyme-linked immunosorbent assay (ELISA) for measuring fecal calprotectin (FC) and C-reactive protein (CRP), and XP-Sysmex for detecting white blood cells. Results: The calprotectin levels below 50 ng/mL were more common in the second age group. The third age group had calprotectin values of 50 ng/mL. In the first age group (25-29 years old), calprotectin frequency was similar. CRP levels were equal in the first age group. In the second age group (30-34 years), negative CRP outnumbered positive CRP. Positive CRP was more common than negative in the third age group of >34 years. The group above 35 had the highest general stool examination (GSE) and antigen Ag detection rates, while those aged 30-34 years in the control group had the highest. Conclusion: Calprotectin levels less than 50 ng/mL were more frequent in the age group of 30-34 years, while calprotectin levels greater than 50 ng/mL were more frequent in the age group <35 years. CRP levels were equal in frequency in the first age group. Negative CRP was more common than positive CRP in the second age group. Positive CRP was more common than negative CRP in the third age group. [ABSTRACT FROM AUTHOR]

Copyright of Saudi Medical Journal is the property of Saudi Medical Journal and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Amebic liver abscesses should be considered in adult males with a liver abscess and a history of travel to endemic areas. Effective treatment includes metronidazole, followed by paromomycin.

Abstract Objective Proteasomes are conserved proteases crucial for proteostasis in eukaryotes and are promising drug targets for protozoan parasites. Yet, the proteasomes of Entamoeba histolytica remain understudied. The study’s objective was to analyse the differences in the substrate binding pockets of amoeba proteasomes from those of host, and computational modelling of β5 catalytic subunit, with the goal of finding selective inhibitors. Results Comparative sequence analysis revealed differences in substrate binding sites of E. histolytica proteasomes, especially in the S1 and S3 pockets of the catalytic beta subunits, implying differences in substrate preference and susceptibility to inhibitors from host proteasomes. This was strongly supported by significantly lower sensitivity to MG132 mediated inhibition of amoebic proteasome β5 subunit’s chymotryptic activity compared to human proteasomes, also reflected in lower sensitivity of E. histolytica to MG132 for inhibition of proliferation. Computational models of β4 and β5 subunits, and a docked β4-β5 model revealed a binding pocket between β4-β5, similar to that of Leishmania tarentolae. Selective inhibitors for visceral leishmaniasis, LXE408 and compound 8, docked well to this pocket. This functional and sequence-based analysis predicts differences between amoebic and host proteasomes that can be utilized to develop rationally designed, selective inhibitors against E. histolytica.

Background: The urease-producing Helicobacter pylori increase the likelihood that pathogenic intestinal protozoa will use the stomach's increased hydrogen potential to propagate the disease. Coinfections exacerbate the onset and severity of gastrointestinal symptoms. This study aimed to assess the prevalence of intestinal parasites/Helicobacter pylori coinfection and contributing factors in patients with gastrointestinal symptoms at Addis Zemen Primary Hospital, Northwest Ethiopia. Methods: From April to July 2023, patients with gastrointestinal problems participated in a cross-sectional study carried out in a hospital. To collect the clinical and sociodemographic data, a questionnaire was employed. Intestinal parasites and Helicobacter pylori were detected using the saline stool wet mount and Helicobacter pylori stool antigen tests, respectively. SPSS version 20 was used to analyze the data and variables with p-values < 0.05 were considered statistically significant. Result: The study included 384 participants in total, of which 47.3% (182/384) were farmers and 50.3% (193/384) were women. Of the study subjects, 69/384 or approximately 18%, had intestinal parasitic infections. In 12% of cases (46/384), Helicobacter pylori were detected. A coinfection of Helicobacter pylori and intestinal parasites was found in 5.5% (21/384) of the subjects. Multiple logistic regression revealed increased risk of coinfection of Helicobacter pylori and intestinal parasites in patients who drink surface water (AOR: 10.7, p = 0.03) family history of Helicobacter pylori (AOR: 3.3, p = 0.024) and those with untrimmed fingers (AOR: 4.9, p = 0.031). Conclusions: Giardia lamblia and Entamoeba histolytica/dispar/ moshkovskii/ bangladeshi complex are the most common protozoans that cause coinfection with Helicobacter pylori. Drinking surface water, family history of Helicobacter pylori and untrimmed fingers are the contributing factors to intestinal parasites/Helicobacter pylori coinfection. [ABSTRACT FROM AUTHOR]

Background: Since 2022, diagnostic laboratories in British Columbia have been advised to replace microscopy for intestinal ova and parasites with infectious diarrhea panel nucleic-acid amplification testing (IDP-NAAT). However, this multiplex assay captures only four common parasites: Cyclospora cayetanensis, Cryptosporidium spp., Entamoeba histolytica, and Giardia spp. Methods: An audit was conducted from September 2022 to August 2023, 1 year prior to implementation of IDP-NAAT in LifeLabs BC, when microscopy was the method used to identify intestinal parasites. Results: Pathogenic parasites were identified in 6149 of 52 221 stool specimens. The most common pathogens were Blastocystis hominis (78.47%), Dientamoeba fragilis (12.23%), Giardia spp. (4.93%), and Cyclospora spp. (1.07%). Entamoeba histolytica/dispar, Strongyloides stercoralis, Cryptosporidium spp., Ascaris lumbricoides, Diphyllobothrium spp., Enterobius vermicularis, Hymenolepis nana, Schistosoma mansoni, Taenia spp., Trichuris trichiura, and Clonorchis sinensis each accounted for less than 1% of the pathogenic parasites identified. Enterobius vermicularis was also identified in 46 of 1569 pinworm paddle specimens. Conclusions: Several potentially pathogenic parasites could be missed if only IDP-NAAT is used to detect intestinal parasites. If indicated, microscopy orders would be needed to capture parasites not detected by IDP-NAAT. [ABSTRACT FROM AUTHOR]

Summary: Purpose: This study was carried out to determine the presence of Entamoeba histolytica in water sources of Niğde province in Turkey, between June and November 2021. Methods: A total of 90 water samples were taken from 15 different water sources (drinking water, well water, spring water, wastewater and dam water) every month and the presence of E. histolytica antigens in the samples was examined by ELISA. Results: The positivity for E. histolytica was determined in 7 (7.7%) of 90 samples. While no antigens were found in any of the samples in June and September, E. histolytica was positive for three samples (20%) in July, one sample (6.6%) in August and October and two samples in November (13.3%). One of 24 dam samples (4.1%), 1 of 12 wastewater samples (8.3%), 1 of 12 well samples (8.3%), and 4 of 24 fountain samples (16.6%) that examined by ELISA were found positive. On the other hand, none of the examined 18 spring samples were positive. In addition, 4 (8.8%) of 45 samples that examined in summer and 3 (6.6%) of 45 samples that examined in autumn were detected positive by using ELISA. Entamoeba histolytica positivity in samples was statistically insignificant in terms of months, water resources and seasons (P > 0.05). Conclusion: As a result, the presence of E. histolytica, which is an important public health problem in water sources, was determined for the first time in Niğde province of Türkiye with this study. [ABSTRACT FROM AUTHOR]

Recently, we published that the monoclonal antibody (D12 mAb) recognizes gp63 of L. mexicana, and it is responsible for COX activity. This D12 mAb exhibited cross-reactivity with Trypanosoma cruzi, Entamoeba histolytica, Acanthamoeba castellanii, and Naegleria fowleri. COX activity assays performed in these parasites suggested the potential presence of such enzymatic activity. In our investigation, we confirmed that wild-type recombinant gp63 exhibits COX-like activity, in contrast to a mutated recombinant gp63 variant. Consequently, our objective was to identify sequences orthologous to gp63 and subsequently analyze the binding of arachidonic acid (AA) to the putative active sites of these proteins. Given the absence of a crystallized structure for this protein in the Protein Data Bank (PDB), it was imperative to first obtain a three-dimensional structure by homology modeling, using leishmanolysin from Leishmania major (PDB ID: LML1) as a template in the Swiss model database. The results obtained through molecular docking simulations revealed the primary interactions of AA close to the Zinc atom present in the catalytic site of gp63-like molecules of several parasites, predominantly mediated by hydrogen bonds with HIS264, HIS268 and HIS334. Furthermore, COX activity was evaluated in commensal species such as E. dispar and during the encystment process of E. invadens. [ABSTRACT FROM AUTHOR]

Entamoeba histolytica is a protozoan parasite belonging to the phylum Amoebozoa that causes amebiasis, a global public health problem. E. histolytica alternates its form between a proliferative trophozoite and a dormant cyst. Trophozoite proliferation is closely associated with amebiasis symptoms and pathogenesis whereas cysts transmit the disease. Drugs are available for clinical use; however, they have issues of adverse effects and dual targeting of disease symptoms and transmission remains to be improved. Development of new drugs is therefore urgently needed. An untargeted lipidomics analysis recently revealed structural uniqueness of the Entamoeba lipidome at different stages of the parasite's life cycle involving very long (26–30 carbons) and/or medium (8–12 carbons) acyl chains linked to glycerophospholipids and sphingolipids. Here, we investigated the physiology of this unique acyl chain diversity in Entamoeba, a non-photosynthetic protist. We characterized E. histolytica fatty acid elongases (EhFAEs), which are typically components of the fatty acid elongation cycle of photosynthetic protists and plants. An approach combining genetics and lipidomics revealed that EhFAEs are involved in the production of medium and very long acyl chains in E. histolytica. This approach also showed that the K3 group herbicides, flufenacet, cafenstrole, and fenoxasulfone, inhibited the production of very long acyl chains, thereby impairing Entamoeba trophozoite proliferation and cyst formation. Importantly, none of these three compounds showed toxicity to a human cell line; therefore, EhFAEs are reasonable targets for developing new anti-amebiasis drugs and these compounds are promising leads for such drugs. Interestingly, in the Amoebazoan lineage, gain and loss of the genes encoding two different types of fatty acid elongase have occurred during evolution, which may be relevant to parasite adaptation. Acyl chain diversity in lipids is therefore a unique and indispensable feature for parasitic adaptation of Entamoeba. Author summary: Entamoeba, a protozoan parasite, thrives in its niche by alternating its form between a proliferative trophozoite and a dormant cyst. Entamoeba parasites were recently shown to generate lipid acyl chain diversity throughout their life cycle. Here, we provide insight into the mechanism generating this acyl chain diversity and its importance in maintaining the parasite's life cycle. We also show that the E. histolytica isozymes responsible for generating this diversity are reasonable targets for developing new anti-amebiasis drugs, and that the herbicides, flufenacet, cafenstrole, and fenoxasulfone, are promising leads for such drugs. Interestingly, gain and loss of the two genes encoding the isozymes controlling acyl chain diversity is plausibly related to E. histolytica host adaptation. [ABSTRACT FROM AUTHOR]

The retromer is a cellular structure that recruits and recycles proteins inside the cell. In mammalian and yeast, the retromer components have been widely studied, but very little in parasites. In yeast, it is formed by a SNX-BAR membrane remodeling heterodimer and the cargo selecting complex (CSC), composed by three proteins. One of them, the Vps26 protein, possesses a flexible and intrinsically disordered region (IDR), that facilitates interactions with other proteins and contributes to the retromer binding to the endosomal membrane. In Entamoeba histolytica, the protozoan parasite responsible for human amoebiasis, the retromer actively participates during the high mobility and phagocytosis of trophozoites, but the molecular details in these events, are almost unknown. Here, we studied the EhVps26 role in phagocytosis. Bioinformatic analyses of EhVps26 revealed a typical arrestin folding structure of the protein, and a long and charged IDR, as described in other systems. EhVps26 molecular dynamics simulations (MDS) allowed us to predict binding pockets for EhVps35, EhSNX3, and a PX domain-containing protein; these pockets were disorganized in a EhVps26 truncated version lacking the IDR. The AlphaFold2 software predicted the interaction of EhVps26 with EhVps35, EhVps29 and EhSNX3, in a model similar to the reported mammalian crystals. By confocal and transmission electron microscopy, EhVps26 was found in the trophozoites plasma membrane, cytosol, endosomes, and Golgi-like apparatus. During phagocytosis, it followed the erythrocytes pathway, probably participating in cargoes selection and recycling. Ehvps26 gene knocking down evidenced that the EhVps26 protein is necessary for efficient phagocytosis. [ABSTRACT FROM AUTHOR]

Objective: In this study; it is aimed to evaluate E. histolytica-specific ELISA adhesin antigen test results in stool samples sent to the medical microbiology laboratory with suspicion of amebiasis. Material and Methods: ELISA (Cellabs, Entamoeba Celisa Path, Brookvale, NSW Australia) adhesin antigen test results, examined on stool samples sent to the medical microbiology laboratory with suspicion of amebiasis in a two-year period between January 2022 and December 2023 were evaluated retrospectively. The data including the test results, gender, age, date and clinical information of the cases of the examined sample were obtained from our hospital’s laboratory data software system. Fisher’s exact chi-square test was used in statistical analyses. Results: Of the 1120 samples in the study, 578 (51.6%) belong to male patients and 542 (48.4%) belong to female patients. The ages of the patients ranged from 0 to 94 years, and the average age was 33.92 (standard deviation: ±24.33). There were 335 (30%) samples from pediatric patients, and the mean age was determined as 4.48 ±4.78. One hundred and sixty-one (14.3%) samples were found positive with the ELISA adhesin antigen test specific to E. histolytica. Of patients with positive test results, 77 (47.8%) were male and 84 (52.2%) were female. Among the positive samples, there were 55 (34.1%) samples from pediatric patients. While there was no statistically significant difference between the positivity status and age and gender of the patients (p>0.05), the seasonal difference was considered significant (p<0.05). Conclusion: In our study, E. histolytica specific ELISA adhesin antigen test positivity was determined as 14.3%, and this rate was found close to the country’s literature data. It has been determined that amoebiasis is more common in tropical and subtropical climates. With the use of ELISA adhesin antigen test specific to E. histolytica, which is cheap, fast and does not require experienced personnel, the rate of misdiagnosis can be reduced and unnecessary treatment applications can be reduced. [ABSTRACT FROM AUTHOR]

Objective: Proteasomes are conserved proteases crucial for proteostasis in eukaryotes and are promising drug targets for protozoan parasites. Yet, the proteasomes of Entamoeba histolytica remain understudied. The study's objective was to analyse the differences in the substrate binding pockets of amoeba proteasomes from those of host, and computational modelling of β5 catalytic subunit, with the goal of finding selective inhibitors. Results: Comparative sequence analysis revealed differences in substrate binding sites of E. histolytica proteasomes, especially in the S1 and S3 pockets of the catalytic beta subunits, implying differences in substrate preference and susceptibility to inhibitors from host proteasomes. This was strongly supported by significantly lower sensitivity to MG132 mediated inhibition of amoebic proteasome β5 subunit's chymotryptic activity compared to human proteasomes, also reflected in lower sensitivity of E. histolytica to MG132 for inhibition of proliferation. Computational models of β4 and β5 subunits, and a docked β4-β5 model revealed a binding pocket between β4-β5, similar to that of Leishmania tarentolae. Selective inhibitors for visceral leishmaniasis, LXE408 and compound 8, docked well to this pocket. This functional and sequence-based analysis predicts differences between amoebic and host proteasomes that can be utilized to develop rationally designed, selective inhibitors against E. histolytica. [ABSTRACT FROM AUTHOR]

The protozoan parasite Entamoeba histolytica is the causative agent of amebiasis, with clinical outcomes ranging from asymptomatic infections to severe invasive diseases. The innate immune system, particularly macrophages, is of paramount importance in resisting the invasion of host tissues and organs by the trophozoites of E. histolytica. Parasite-derived pathogenic factors, such as lectins, play a pivotal role in the promotion of macrophage polarization phenotypes that have undergone alteration. Nevertheless, the precise mechanisms by which E. histolytica modulates immune polarization remain largely unknown. The current study focused on the immunomodulatory effects of the Igl-C fragment of E. histolytica Gal/GalNAc lectin on macrophage polarization. These results demonstrated that Igl-C could induce the secretion of IL-1b, IL-6, and other cytokines, activating a mixed M1/M2 polarization state. M1 polarization of macrophages occurs in the early stages and gradually transitions to M2 polarization in the later stages, which may contribute to the persistence of the infection. Igl-C induces the macrophage M1 phenotype and causes the release of immune effector molecules, including iNOS and cytokines, by activating the NF-kB p65 and JAK-STAT1 transcription factor signaling pathways. Furthermore, Igl-C supports the macrophage M2 phenotype via JAK-STAT3 and IL-4-STAT6 pathways, which activate arginase expression in later stages, contributing to the tissue regeneration and persistence of the parasite. The involvement of distinct signaling pathways in mediating this response highlights the complex interplay between the parasite and the host immune system. These findings enhance our understanding of the Igl-Cmediated pathogenic mechanisms during E. histolytica infection. [ABSTRACT FROM AUTHOR]

Peptidyl-prolyl cis/trans isomerases (PPIases) are present in a wide variety of microorganisms, including protozoan parasites such as Trypanosoma cruzi, Trypanosoma brucei, Trichomonas vaginalis, Leishmania major, Leishmania donovani, Plasmodium falciparum, Plasmodium vivax, Entamoeba histolytica, Giardia intestinalis, Cryptosporidium parvum, and Cryptosporidium hominis, all of which cause important neglected diseases. PPIases are classified as cyclophilins, FKBPs, or parvulins and play crucial roles in catalyzing the cis-trans isomerization of the peptide bond preceding a proline residue. This activity assists in correct protein folding. However, experimentally, the biological structure–function characterization of PPIases from these protozoan parasites has been poorly addressed. The recombinant production of these enzymes is highly relevant for this ongoing research. Thus, this review explores the structural diversity, functions, recombinant production, activity, and inhibition of protozoan PPIases. We also highlight their potential as biotechnological tools for the in vitro refolding of other recombinant proteins from these parasites. These applications are invaluable for the development of diagnostic and therapeutic tools. [ABSTRACT FROM AUTHOR]

Cell migration modes can vary, depending on a number of environmental and intracellular factors. The high motility of the pathogenic amoeba Entamoeba histolytica is a decisive factor in its ability to cross the human colonic barrier. We used quantitative live imaging techniques to study the migration of this parasite on fibronectin, a key tissue component. Entamoeba histolytica amoebae on fibronectin contain abundant podosome-like structures. By using a laminar flow chamber, we determined that the adhesion forces generated on fibronectin were twice those on non-coated glass. When migrating on fibronectin, elongated amoeboid cells converted into fan-shaped cells characterized by the presence of a dorsal column of F-actin and a broad cytoplasmic extension at the front. The fan shape depended on the Arp2/3 complex, and the amoebae moved laterally and more slowly. Intracellular measurements of physical variables related to fluid dynamics revealed that cytoplasmic pressure gradients were weaker within fan-shaped cells; hence, actomyosin motors might be less involved in driving the cell body forward. We also found that the Rho-associated coiled-coil containing protein kinase regulated podosome dynamics. We conclude that E. histolytica spontaneously changes its migration mode as a function of the substrate composition. This adaptive ability might favour E. histolytica's invasion of human colonic tissue. By combining microfluidic experiments, mechanical modelling, and image analysis, our work also introduces a computational pipeline for the study of cell migration. Author summary: Even though we know increasingly well how cells move in tissues, the question of whether parasites modify their migration as a function of the host tissue environment requires further investigation. Cell motility is triggered by environmental stimuli, and the migration mode depends on adhesion, activated enzymatic pathways, and changes in the actin cytoskeleton dynamics and thus cell morphology. The protozoan intestinal parasitic Entamoeba histolytica is the causative agent of amoebiasis (a major food- and water-borne disease) in humans. During invasion of the colon, E. histolytica degrades the mucus layer, adheres to and removes the epithelium, progresses along the fibronectin-rich basement membrane towards the crypts of Lieberkühn, and penetrates the mucosa. This invasive process raises the question of how the parasite moves in a varying intestinal environment. E. histolytica is known to display an amoeboid migration mode and binds fibronectin through podosome adhesion structures. We hypothesized that fibronectin triggers signalling pathways that influence the trophozoite's migration. We found that E. histolytica can migrate with a non-amoeboid mode on fibronectin-coated glass. Our finding suggests that for tissue invasion, E. histolytica's migration mode is modulated to suit the environment. [ABSTRACT FROM AUTHOR]

Entamoeba histolytica is the protozoan causative of human amoebiasis. The EhADH adhesin (687 aa) is a protein involved in tissue invasion, phagocytosis and host-cell lysis. EhADH adheres to the prey and follows its arrival to the multivesicular bodies. It is an accessory protein of the endosomal sorting complexes required for transport (ESCRT) machinery. Here, to study the role of different parts of EhADH during virulence events, we produced trophozoites overexpressing the three domains of EhADH, Bro1 (1–400 aa), Linker (246–446 aa) and Adh (444–687 aa) to evaluate their role in virulence. The TrophozBro11–400 slightly increased adherence and phagocytosis, but these trophozoites showed a higher ability to destroy cell monolayers, augment the permeability of cultured epithelial cells and mouse colon, and produce more damage to hamster livers. The TrophozLinker226–446 also increased the virulence properties, but with lower effect than the TrophozBro11–400. In addition, this fragment participates in cholesterol transport and GTPase binding. Interestingly, the TrophozAdh444–687 produced the highest effect on adherence and phagocytosis, but it poorly influenced the monolayers destruction; nevertheless, they augmented the colon and liver damage. To identify the protein partners of each domain, we used recombinant peptides. Pull-down assays and mass spectrometry showed that Bro1 domain interplays with EhADH, Gal/GalNAc lectin, EhCPs, ESCRT machinery components and cytoskeleton proteins. While EhADH, ubiquitin, EhRabB, EhNPC1 and EhHSP70 were associated to the Linker domain, and EhADH, EhHSP70, EhPrx and metabolic enzymes interacted to the Adh domain. The diverse protein association confirms that EhADH is a versatile molecule with multiple functions probably given by its capacity to form distinct molecular complexes. [ABSTRACT FROM AUTHOR]

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OBJECTIVE: To investigate the prevalence of parasitic infections in the human population and identify the related risk factors of Gonabad city, southeast of Khorasan Razavi province, northeastern Iran. METHODOLOGY: This cross-sectional study was performed from March to April 2019 in Gonabad city in Khorasan Razavi province in northeastern Iran. One thousand thirty-eight stool samples were examined using parasitological methods like direct wet-mounting, formalin-ether concentration and Gomori's trichrome staining. Moreover, the socio-demographic data of inhabitants were recorded. All statistical analyses were performed using SPSS software (version 16). Qualitative variables were reported as numbers and percentages. RESULTS: The overall intestinal parasitic infection prevalence was 18.1 %, whereas this value for protozoan agents was 16.7% (95% CI 14.14 to 19) and 1.3% (95% CI 0.7 to 2) for helminth parasites. The highest parasitic agent prevalence belongs to Blastocystis sp (12.4 %), Entamoeba coli (1.8%) and Enterobius vermicularis (1.2%), respectively. Giardia lamblia (1.1%) Entamoeba histolytica/dispar (0.3%), Iodamoeba butschlii (0.3%), Chilomastix mesnili (0.3%), Hymenolepis nana (0.2 %) Dientamoeba fragilis (0.2%) and Trichomonas hominis (0.3%) were other identified parasites. Multivariate logistic regression analysis revealed a significant association of parasitic intestinal infections with factors such as age, residency status and source of drinking water (P<0.05). CONCLUSION: Our findings approved that the protozoan agents have a relatively high prevalence among the studied population. So, it could be severe enough to alarm the entities implementing healthcare services measures for parasitic intestinal infection control. [ABSTRACT FROM AUTHOR]

This case report provides data on unique challenges related to amoebiasis diagnostics and treatment in non-endemic regions. The presented case report is focused a 28-year-old male patient of Indian origin, temporarily living in Poland, who was diagnosed with an amoebic liver abscess. The patient presented with a range of non-specific symptoms including shortness of breath, chest pain, and fever. The differential diagnosis included cardio-pulmonary diseases, a range of tropical diseases such as malaria or typhoid fever, bacterial abscesses, and malignancies, necessitating a comprehensive, multi-modal diagnostic approach. This approach included an extensive review of patient history, physical examination, and various laboratory and imaging investigations. A further challenge in this case was the unavailability of standard cysticidal treatments in Poland, which required individualized therapeutic strategy. Despite these obstacles, the patient was successfully treated using an alternative regimen of intravenous metronidazole, ceftriaxone, doxycycline, chloroquine, and finally, trimethoprim/sulfamethoxazole (treatment with metronidazole was used as a base drug, due to the lack of typical cysticidal treatment, an alternative treatment was added: chloroquine is a recommended drug used in the treatment of pregnant patients, in addition, doxycycline showed in vitro activity against Entamoeba histolytica). This therapeutic journey underscored the value of adaptability in treatment protocols, particularly in regions where certain resources may not be readily available. This case report underlines the importance of broadening the differential diagnosis in non-endemic regions to include tropical diseases, particularly in the context of increasing global travel and migration. It also highlights the significance of employing comprehensive diagnostic strategies and adaptable treatment protocols in such scenarios. In addition, the report reiterates the need for global collaboration and education among healthcare providers to effectively manage tropical diseases, especially in non-endemic regions. Through its exploration of the complexities associated with diagnosing and managing amebiasis in a non-endemic region, this report offers valuable insights to clinicians worldwide. [ABSTRACT FROM AUTHOR]

During intestinal and liver invasion by the protozoan parasite Entamoeba histolytica, extensive tissue destruction linked to large neutrophil infiltrates is observed. It has been proposed that microbicidal components of neutrophils are responsible for the damage, however, the mechanism by which they are released and act in the extracellular space remains unknown. In previous studies, we have shown that E. histolytica trophozoites induce NET formation, leading to the release of neutrophil granule content into extruded DNA. In this work, we evaluate the possible participation of NETs in the development of amoeba-associated pathology and analyze the contribution of anti-microbial components of the associated granules. E. histolytica-induced NETs were isolated and their effect on the viability and integrity of HCT 116 colonic and Hep G2 liver cultures were evaluated. The results showed that simple incubation of cell monolayers with purified NETs for 24 h resulted in cell detachment and death in a dose-dependent manner. The effect was thermolabile and correlated with the amount of DNA and protein present in NETs. Pretreatment of NETs with specific inhibitors of some microbicidal components suggested that serine proteases, are mostly responsible for the damage caused by NETs on HCT 116 cells, while the MPO activity was the most related to Hep G2 cells damage. Our study also points to a very important role of DNA as a scaffold for the activity of these proteins. We show evidence of the development of NETs in amoebic liver abscesses in hamsters as a preamble to evaluate their participation in tissue damage. In conclusion, these studies demonstrate that amoebic-induced NETs have potent cytotoxic effects on target cells and, therefore, may be responsible for the intense damage associated with tissue invasion by this parasite.

In this study, n-butyl and iso-butyl quinoxaline-7-carboxylate-1,4-di-N-oxide derivatives were evaluated in vitro against Giardia lamblia (G. lamblia), Trichomonas vaginalis (T. vaginalis), and Entamoeba histolytica (E. histolytica). The potential mechanism of action determination was approached by in silico analysis on G. lamblia and T. vaginalis triosephosphate isomerase (GlTIM and TvTIM, respectively), and on E. histolytica thioredoxin reductase (EhTrxR). Enzyme inactivation assays were performed on recombinant GlTIM and EhTrxR. Compound T-167 showed the best giardicidal activity (IC50 = 25.53 nM) and the highest inactivation efficiency against GlTIM without significantly perturbing its human homolog. Compounds T-142 and T-143 showed the best amoebicidal (IC50 = 9.20 nM) and trichomonacidal (IC50 = 45.20 nM) activity, respectively. Additionally, T-143 had a high activity as giardicial (IC50 = 29.13 nM) and amoebicidal (IC50 = 15.14 nM), proposing it as a broad-spectrum antiparasitic agent. Compounds T-145, and T-161 were the best EhTrxR inhibitors with IC50 of 16 µM, and 18 µM, respectively.

Entamoeba histolytica, the causative agent of amebiasis, is one of the top three parasitic causes of mortality worldwide. However, no vaccine exists against amebiasis. Using a lead candidate vaccine containing the LecA fragment of Gal-lectin and GLA-3M-052 liposome adjuvant, we immunized rhesus macaques via intranasal or intramuscular routes. The vaccine elicited high-avidity functional humoral responses as seen by the inhibition of amebic attachment to mammalian target cells by plasma and stool antibodies. Importantly, antigen-specific IFN-γ-secreting peripheral blood mononuclear cells (PBMCs) and IgG/IgA memory B cells (BMEM) were detected in immunized animals. Furthermore, antigen-specific antibody and cellular responses were maintained for at least 8 months after the final immunization as observed by robust LecA-specific BMEM as well as IFN-γ+ PBMC responses. Overall, both intranasal and intramuscular immunizations elicited a durable and functional response in systemic and mucosal compartments, which supports advancing the LecA+GLA-3M-052 liposome vaccine candidate to clinical testing.

ABSTRACT Entamoeba histolytica is an enteric protozoan parasite that causes human amebic colitis and extraintestinal abscesses. As a prerequisite for parasite colonization and invasion, adherence of E. histolytica is predominantly mediated by galactose (Gal)- and N-acetyl-d-galactosamine (GalNAc)-inhibitable lectins. The intermediate subunit (Igl) of Gal-/GalNAc-inhibitable lectin is a cysteine-rich protein containing multiple CXXC motifs and is considered a key factor affecting trophozoite’s pathogenicity. However, details of the function of Igl during parasite adherence remain unclear. Here, using segmentally expressed Igl proteins and a CHO cell model transfected with Igl fragments, we identified a carbohydrate-recognition domain (CRD)-like region between amino acids 989 and 1,088. Through single- and double-point mutations in the Igl segments, two core CXXC motifs responsible for carbohydrate recognition in the CRD-like region, which are highly conserved among several lectins, were confirmed. In addition to adhesion, the roles of CRD-like region and its core CXXC motifs in various pathogenic effects were further explored. To our knowledge, this is the first report showing an adhesion-related region in E. histolytica Igl. The identification and characterization of this CRD-like region provides further insights into molecular mechanisms underlying E. histolytica pathogenicity and also aids in the determination of a potential drug target in this parasite.IMPORTANCEEntamoeba histolytica adhesion mainly depends on galactose (Gal)-/N-acetyl-d-galactosamine (GalNAc)-inhibitable lectins, subsequently triggering a series of amebic reactions. Among the three subunits of Gal-/GalNAc-inhibitable lectin, heavy subunit and intermediate subunit (Igl) have exhibited lectin activity, but that of Igl remains poorly understood. In this study, we confirmed a carbohydrate-recognition domain (CRD)-like limiting region in E. histolytica Igl and further identified its two core CXXC motifs responsible for carbohydrate recognition. Moreover, the role of Igl's CRD-like region and its CXXC motifs in hemolysis and pathogenic effects was explored. This is the first study to determine an adhesion-related region in E. histolytica Igl protein, providing a new reference direction for subsequent research studies. Since the potential homogeneity of galectin-2 in several mammals and Igl CRD-like region, it could be meaningful to relate the corresponding pathogeneses and phenotypes of these two proteins. Except for adhesion, studies on the involvement of Igl CRD-like region in different parasite–host interactions are also promising.

Worm and parasitic infestation is a common problem in children with most occurring in school age . Low socio-conomic status and poor sanitary hygiene has been associated with worm infesetation. Though it is rare in infancy, there had been reports in these group of children. However there has been no documented case of multiple infestation in infancy. We hereby report an infant with recurrent anemia and melena and bloody stools caused by anklylostomaduodenale and entamoebahistolytica

This narrative review was aimed at collecting updated knowledge on the risk factors, illnesses caused, and measures for the prevention of protozoan infections transmitted by food and drinking water. Reports screened dated from 2019 to the present and regarded global prevalence in food handlers, occurrence in food and drinking water, impact on human health, and recently reported outbreaks and cases of severe infections attributable to the dietary route. Cryptosporidium spp., Cyclospora cayetanensis, Entamoeba histolytica, and Cystoisospora belli were the protozoans most frequently involved in recently reported waterborne and foodborne outbreaks and cases. Blastocystis hominis was reported to be the most widespread intestinal protozoan in humans, and two case reports indicated its pathogenic potential. Dientamoeba fragilis, Endolimax nana, and Pentatrichomonas hominis are also frequent but still require further investigation on their ability to cause illness. A progressive improvement in surveillance of protozoan infections and infection sources took place in developed countries where the implementation of reporting systems and the application of molecular diagnostic methods led to an enhanced capacity to identify epidemiological links and improve the prevention of foodborne and waterborne protozoan infections. [ABSTRACT FROM AUTHOR]

Background: Enterocolitis and gastroenteritis remain major health problems, particularly in children living in developing countries. Intestinal protozoa, such as Entamoeba histolytica, Blastocystis, and Cyclospora, are frequently associated with these conditions. Amebic colitis can cause serious complications, including fulminant necrotizing colitis, toxic megacolon, extraintestinal amebiasis, and stunting in children. The diagnosis of amoebiasis is challenging, relying on microscopic examination, which cannot distinguish E. histolytica from the nonpathogenic E. dispar and E. moshkovskii. Therefore, this study aimed to identify intestinal parasites, particularly Entamoeba, their prevalence, and the clinical characteristics of patients admitted for enterocolitis and gastroenteritis at a tertiary-referral hospital. Material and Methods: A cross-sectional, retrospective study was conducted at a national, tertiary-referral government hospital, in Jakarta. Of the 111 retrieved medical records from hospitalized patients with enterocolitis and gastroenteritis, for which parasitology feces were examined, 54 fecal samples (48.6%) were still available in the parasitology laboratory storage. All fecal samples underwent the following tests: 1) direct stool examination, after staining with 1% Lugol’s solution, and using the water–ether concentration method; 2) modified acid-fast staining for coccidian parasites; 3) Jones’ culture medium to detect Blastocystis; 4) copro-antigen assay to detect Cryptosporidium and Giardia; and 5) a polymerase chain reaction (PCR) assay to identify Entamoeba. Clinical and demographic data were obtained from the medical records. Results: Largely, patients (44.1%) were from the cohort of young children ≤5 years old, followed by adults aged 19–60 years old (24.3%). Both cohorts exhibited polyparasitism. Intestinal parasites were detected in 17 out of the 54 samples (31.4%). These included 6 (11.1%), 2 (3.7%),5 (9.2%), 3 (5.5%), 2 (3.7%), and 1 (1.8%) samples that were positive for Blastocystis, E dispar, E. histolytica, E. moshkovskii, Cryptosporidium, and Dientamoeba fragilis, respectively. PCR analysis revealed that 10 samples were positive for Entamoeba infection, eight of which originated from pediatric patients. Conclusion: At a national tertiary-referral hospital in Indonesia, Entamoeba infection was the most prevalent parasite among pediatric patients with enterocolitis. E. histolytica and E. moshkovskii were the two main species identified by PCR. Therefore, PCR assays and fecal occult–blood tests are recommended in cases of enterocolitis and gastroenteritis. [ABSTRACT FROM AUTHOR]

In humans, Entamoeba histolytica is the main pathogen causing various amoebiases, while E. moshkovskii falls between being a pathogen and non-pathogen. The two species have similar behavior patterns but differ significantly in pathogenicity, with previous studies and clinical data indicating that E. moshkovskii has a low level of pathogenicity. Meaningfully, the biological characteristics of E. moshkovskii make it a potential model organism and a protein display platform for studying the functions of important Entamoeba proteins. Here, an Amoeba-pcDNA3.1 vector capable of overexpressing E. histolytica-sourced Igl-C protein was constructed and successfully transfected into E. moshkovskii. High levels of expression of the Igl-C, EGFP, and NeoR genes were identified in Igl-C-transfected trophozoites using qRT-PCR, and they were subsequently confirmed using immunoblotting. Transfection of Igl-C protein improved the adherence and phagocytosis of E. moshkovskii, demonstrating that E. histolytica Igl mediated amoebic adhesion. Moreover, as a manifestation of protein virulence, the ability of post-transfected trophozoites to induce inflammation in host macrophages was also enhanced. In conclusion, this study utilizing the characteristics of E. moshkovskii confirmed its potential to serve as a model organism. E. moshkovskii could replace E. histolytica as the target of gene editing, allowing more efficient study of amoebic pathogenicity. [ABSTRACT FROM AUTHOR]

Entamoeba histolytica is a protozoan parasite that causes amoebiasis, an infection primarily affecting the intestines. The study aimed to determine the phylogenetic placement of six isolates of E. histolytica that were amplified from three genetic loci, rRNA, cysteine proteinase, and amoebapore C to evaluate the genetic variation pattern of each sequence. DNA extracted from cultured and frozen stool samples using PrestoTM Stool DNA Extraction Kit and Geneaid kits was analyzed for E. histolytica diversity in amoebiasis patients at Baquba General Hospital from March-June 2023. Polymerase chaing kreaction (PCR) and sequencing of virulence genes (rRNA, cysteine proteinase, amoebapore C) followed by phylogenetic analysis revealed genetic variation and potential implications for pathogenicity. The conducted sequencing reactions confirmed the precise identification of the examined samples, which were determined to be attributed to variable isolates of E. histolytica. Aligning the rRNA sequences of samples A1 and A2 with the most closely related rRNA sequences of E. histolytica (GenBank acc. no. OP925909.1) revealed the presence of two nucleic acid variants (212C>T in A1, and 427G>C in A1 and A2) not present in the reference sequences. Aligning the cysteine proteinase sequences of samples B1 and B2 with the most closely related genomic DNA sequences of E. histolytica (GenBank acc. no. X91642.1) revealed the presence of four nucleic acid variants (95A>T and 96A>T in B1 and B2, and 246A>T and 247A>T in B2) not present in the reference sequences. Translation of nucleic acid sequences to amino acid residues showed that both 95A>T and 96A>T variants caused a missense effect of p.46E>V, while both 246A>T and 247A>T variants showed another missense effect of p.97T>S. Aligning the amoebapore C sequences of samples C1 and C2 with the most closely related genomic sequences of E. histolytica (GenBank acc. no. X76903.1) revealed their entire similarity. It was inferred from the tree that the investigated protozoan samples occupied distinct phylogenetic positions and were suitable for the vicinity of various clinical isolates that were isolated from several locations worldwide. However, the rRNA-based tree provided ample genetic diversity with more effective discrimination compared with the other used loci. [ABSTRACT FROM AUTHOR]