Amoebicidal Effect Research Articles

Amoebicidal Effect of COVID Box Molecules against Acanthamoeba: A Study of Cell Death.

Acanthamoeba spp. can cause a sight threatening disease. At present, the current treatments used to treat Acanthamoeba spp. Infections, such as biguanide-based antimicrobials, remain inefficacious, with the appearance of resistant forms and high cytotoxicity to host cells. In this study, an initial screening was conducted against Acanthamoeba castellanii Neff and murine macrophages J774A.1 using alamarBlue™. Among the 160 compounds included in the cited box, 90% exhibited an inhibition of the parasite above 80%, while only 18.75% of the compounds inhibited the parasite with a lethality towards murine macrophage lower than 20%. Based on the amoebicidal activity, the cytotoxicity assay, and availability, Terconazole was chosen for the elucidation of the action mode in two clinical strains, Acanthamoeba culbertsoni and Acanthamoeba castellanii L10. A fluorescence image-based system and proteomic techniques were used to investigate the effect of the present azole on the cytoskeleton network and various programmed cell death features, including chromatin condensation and mitochondria dysfunction. Taking all the results together, we can suggest that Terconazole can induce programmed cell death (PCD) via the inhibition of sterol biosynthesis inhibition.

Modelling amoebic brain infection caused by Balamuthia mandrillaris using a human cerebral organoid.

The lack of disease models adequately resembling human tissue has hindered our understanding of amoebic brain infection. Three-dimensional structured organoids provide a microenvironment similar to human tissue. This study demonstrates the use of cerebral organoids to model a rare brain infection caused by the highly lethal amoeba Balamuthia mandrillaris. Cerebral organoids were generated from human pluripotent stem cells and infected with clinically isolated B. mandrillaris trophozoites. Histological examination showed amoebic invasion and neuron damage following coculture with the trophozoites. The transcript profile suggested an alteration in neuron growth and a proinflammatory response. The release of intracellular proteins specific to neuronal bodies and astrocytes was detected at higher levels postinfection. The amoebicidal effect of the repurposed drug nitroxoline was examined using the human cerebral organoids. Overall, the use of human cerebral organoids was important for understanding the mechanism of amoeba pathogenicity, identify biomarkers for brain injury, and in the testing of a potential amoebicidal drug in a context similar to the human brain.

Amoebicidal effect of chlorine dioxide gas against pathogenic Naegleria fowleri and Acanthamoeba polyphaga.

The pathogenic free-living amoebae, Naegleria fowleri and Acanthamoeba polyphaga, are found in freshwater, soil, and unchlorinated or minimally chlorinated swimming pools. N. fowleri and A. polyphaga are becoming problematic as water leisure activities and drinking water are sources of infection. Chlorine dioxide (ClO2) gas is a potent disinfectant that is relatively harmless to humans at the concentration used for disinfection. In this study, we examined the amoebicidal effects of ClO2 gas on N. fowleri and A. polyphaga. These amoebae were exposed to ClO2 gas from a ready-to-use product (0.36 ppmv/h) for 12, 24, 36, and 48h. Microscopic examination showed that the viability of N. fowleri and A. polyphaga was effectively inhibited by treatment with ClO2 gas in a time-dependent manner. The growth of N. fowleri and A. polyphaga exposed to ClO2 gas for 36h was completely inhibited. In both cases, the mRNA levels of their respective actin genes were significantly reduced following treatment with ClO2 gas. ClO2 gas has an amoebicidal effect on N. fowleri and A. polyphaga. Therefore, ClO2 gas has been proposed as an effective agent for the prevention and control of pathogenic free-living amoeba contamination.

Chitosan nanoparticles improve the effectivity of miltefosine against Acanthamoeba.

Acanthamoeba keratitis (AK) is a corneal sight-threatening infection caused by the free-living amoebae of the genus Acanthamoeba. Early and appropriate treatment significantly impacts visual outcomes. Mucoadhesive polymers such as chitosan are a potential strategy to prolong the residence time and bioavailability of the encapsulated drugs in the cornea. Regarding the recent administration of miltefosine (MF) for treating resistant AK, in the present study, we synthesized miltefosine-loaded chitosan nanoparticles (MF-CS-NPs) and evaluated them against Acanthamoeba. Chitosan nanoparticles (CNPs) were prepared using the ionic gelation method with negatively charged tripolyphosphate (TPP). The zeta-potential (ZP) and the particle size of MF-CS-NPs were 21.8±3.2 mV and 46.61±18.16 nm, respectively. The release profile of MF-CS-NPs indicated linearity with sustained drug release. The cytotoxicity of MF-CS-NPs on the Vero cell line was 2.67 and 1.64 times lower than free MF at 24 and 48 hours. This formulation exhibited no hemolytic activity in vitro and ocular irritation in rabbit eyes. The IC50 of MF-CS-NPs showed a significant reduction by 2.06 and 1.69-fold in trophozoites at 24 and 48 hours compared to free MF. Also, the MF-CS-NPs IC50 in the cysts form was slightly decreased by 1.26 and 1.21-fold at 24 and 48 hours compared to free MF. The MF-CS-NPs were more effective against the trophozoites and cysts than free MF. The nano-chitosan formulation was more effective on trophozoites than the cysts form. MF-CS-NPs reduced toxicity and improved the amoebicidal effect of MF. Nano-chitosan could be an ideal carrier that decreases the cytotoxicity of miltefosine. Further analysis in animal settings is needed to evaluate this nano-formulation for clinical ocular drug delivery.

Amoebicidal effect of synthetic indoles against Acanthamoeba spp.: a study of cell death.

Organic and synthetic chemistry plays a crucial role in drug discovery fields. Moreover, chemical modifications of available molecules to enhance their efficacy, selectivity and safety have been considered as an attractive approach for the development of new bioactive agents. Indoles, a versatile group of natural heterocyclic compounds, have been widely used in pharmaceutical industry due to their broad spectrum of activities including antimicrobial, antitumoral and anti-inflammatory among others. Herein, we report the amoebicidal activity of different indole analogs on Acanthamoeba castellanii Neff. Among the 40 tested derivatives, eight molecules were able to inhibit this protistan parasite. The structure-activity relationship (SAR) analysis of their anti-Acanthamoeba activity would suggest that a carboxylation of C-3 position and the incorporation of halogen as chlorine/fluorine would enhance their biological profile, presumably by increasing their lipophilicity and therefore their ability to cross the cell membrane. Fluorescence image base system was used to investigate the effect of indole 6o c-6 on the cytoskeleton network and various programmed cell death features. We were able to highlight that the methyl 6-chloro-1H-indole-3-carboxylate could induce program cell death by the mitochondrial dysfunction.

Lactase can target cellular differentiation of Acanthamoeba castellanii belonging to the T4 genotype.

The free living Acanthamoeba spp. are ubiquitous amoebae associated with potentially blinding disease known as Acanthamoeba keratitis (AK) and a fatal central nervous system infection granulomatous amoebic encephalitis (GAE). With the inherent ability of cellular differentiation, it can phenotypically transform to a dormant cyst form from an active trophozoite form. Acanthamoeba cysts are highly resistant to therapeutic agents as well as contact lens cleaning solutions. One way to tackle drug resistance against Acanthamoeba is by inhibiting the formation of cysts from trophozoites. The biochemical analysis showed that the major component of Acanthamoeba cyst wall is composed of carbohydrate moieties such as galactose and glucose. The disaccharide of galactose and glucose is lactose. In this study, we analyzed the potential of lactase enzyme to target carbohydrate moieties of cyst walls. Amoebicidal assessment showed that lactase was ineffective against trophozoite of A. castellanii but enhanced amoebicidal effects of chlorhexidine. The lactase enzyme did not show any toxicity against normal human keratinocyte cells (HaCaT) at the tested range. Hence, lactase can be used for further assessment for development of potential therapeutic agents in the management of Acanthamoeba infection as well as formulation of effective contact lens disinfectants.

The Potential Amoebicidal Effect of Some Milk forms as an Inhibitor for Entamoeba histolytica Viability Isolated from Diarrhea

Milk includes numerous compounds such as lacto-peroxidase, lactoferrin, immunoglobulin G, secretory immunoglobulin A, and Lysozymes The invasive trophozoites of E. histolytica are capable to inhabit host tissues and provoke diseases such as intestinal colitis. The objective of the study was to compare different forms of milk’s effect on E. histolytica trophopzoite by measuring protein concentration in vitro using a spectrophotometer and number of trophozoite by microscopy. For this purpose, E. histolytica trophozoites were exposed to several types of milk (human, sheep, cattle, goats, and formula) at concentrations of 6, 12, 18, and 24% for each type after 8, 16, and 24 hours of incubation. According to the findings, sheep and goat milk had lower total protein concentrations after 8 hours than other types of milk when compared to the control group. After 16 hours of incubation, raise the milk concentration to 24% leading to a reduction in the formula milk's total protein concentration to 5g/l. after 8 and 16 hours of incubation, sheep milk and cattle have significantly lower total protein concentrations (24% vs. 8%), respectively. The action of formula and human milk is increased by 24-hour incubation. Increase the trophozoite inhibition percentage compared to metronidazole in human milk, which was followed by sheep and cattle milk with respective values of 60.9%, 56.1%, and 46.3% respectively. Human milk, goat and formula milk had the greatest inhibitory activity after incubation 24hrs. at 24%.the extent of amoebicidal activity of milk was shown to depend on incubation period and concentration of milk.

The anti-amoebic potential of carboxamide derivatives containing sulfonyl or sulfamoyl moieties against brain-eating Naegleria fowleri.

Naegleria fowleri is a free-living thermophilic flagellate amoeba that causes a rare but life-threatening infection called primary amoebic meningoencephalitis (PAM), with a very high fatality rate. Herein, the anti-amoebic potential of carboxamide derivatives possessing sulfonyl or sulfamoyl moiety was assessed against pathogenic N. fowleri using amoebicidal, cytotoxicity and cytopathogenicity assays. The results from amoebicidal experiments showed that derivatives dramatically reduced N. fowleri viability. Selected derivatives demonstrated IC50 values at lower concentrations; 1j showed IC50 at 24.65 μM, while 1k inhibited 50% amoebae growth at 23.31 μM. Compounds with significant amoebicidal effects demonstrated limited cytotoxicity against human cerebral microvascular endothelial cells. Finally, some derivatives mitigated N. fowleri-instigated host cell death. Ultimately, this study demonstrated that 1j and 1k exhibited potent anti-amoebic activity and ought to be looked at in future studies for the development of therapeutic anti-amoebic pharmaceuticals. Further investigation is required to determine the clinical relevance of our findings.

Amoebicidal effect of Fluconazole and Verapamil together against trophozoites and cysts of Acanthamoeba castellanii

Acanthamoeba species are an important pathogen that causes Acanthamoeba keratitis, which causes a visual loss, and encephalitis, which lead to the central nervous system infection and death. The options available to successfully treat Acanthamoeba infections have limited success. New therapeutic approaches must thus be developed, and especially combination medication therapy may be a successful and an effective strategy. The aim of this study was to assess the combination efficacy of verapamil and fluconazole against Acanthamoeba trophozoites and cysts. The effects of drugs on growth inhibition against Acanthamoeba were tested using amoebicidal assays. The viability of Acanthamoeba was assessed using Trypan blue and hemocytometer counts. The effect of three different concentrations of fluconazole, verapamil and fluconazole + verapamil combination on growth inhibition against Acanthamoeba trophozoites and cysts was significant compared to the control (p

In Vitro Stability of the Biological Activity of Voriconazole against Acanthamoeba castellanii

Acanthamoeba keratitis (AK) is a rare cornea disease caused by species of the Acanthamoeba genus. The antifungal voriconazole blocks the ergosterol synthesis in the protozoan membrane and is active against the cysts and trophozoites of Acanthamoeba spp. Due to the low stability of voriconazole, its options for eye drops are scarce. This study aimed to investigate the stability of the biological activity of voriconazole against two strains of Acanthamoeba castellanii and one clinical isolate from a patient with AK. To evaluate the stability of the biological activity of voriconazole, strains of A. castellanii (ATCC 50492) were exposed to different periods and voriconazole concentrations stored at 4 °C for 7, 15, and 30 days. The cytotoxicity assays were performed using SIRC (ATCC CCL-60™) cell line. The results indicated the amoebicidal effect of voriconazole against Acanthamoeba spp. within 24 h and 48 h of exposure, and the voriconazole solution was stable and retained antiamoebic activity when stored at 4 °C for up to 30 days. In the cytotoxicity test, the result demonstrated low cytotoxicity of the drug to the corneal rabbit cell line. However, there is a need to carry out further synergistic effects with other antiamoebic drugs and then in vivo experiments in the AK animal model.

Amoebicidal and Cytotoxic Activity of Propolis collected from Different Regions in Turkey on Acanthamoeba castellanii Trophozoites

Objective: The present study aimed to investigate the amoebicidal effects of propolis collected from different regions in Turkey on Acanthamoeba trophozoites. Method: The propolis was collected from different geographic sites (Van, Erzurum, Gümüşhane, Ordu, Rize, and Muğla) in Turkey. Different concentrations of propolis ethanolic extract (in quantities from 1, 2, 3, 4, 5, 6, 7, and 8 mg/mL) and the same volume of trophozoites in 100 μl culture were blended for the identification of the amoebicidal efficiency of propolis. Results: The growth of trophozoites stopped in Turkish propolis extracts with 50% inhibitory concentrations (IC50)/48h for 5 mg/mL extract solution. Propolis showed more potent inhibitory effects on Acanthamoeba trophozoites at concentrations of 7, 6, 5, 4, and 3 mg/mL for 72 h. Propolis extract substantially inhibited human bronchial epithelial cells, especially at higher concentrations (7, 8, and 16 mg/mL). Propolis can kill Acanthamoeba trophozoites at a concentration (of 3-6 mg/mL) but is safe for human bronchial epithelial cells at the same concentrations after 72 h treatment, this paves the way for propolis to be an alternative source of therapeutic drugs in the treatment of Acanthamoeba spp. Conclusion: Acanthamoeba infection still cannot be treated with drugs. In this study, propolis collected from different regions of Turkey showed amoebic and cytotoxic activities. Propolis extract, which is a natural product that can be used against Acanthamoeba trophozoites can be an alternative source of therapeutic drugs in the treatment of Acanthamoeba, supported by in vivo studies.

Anti-Balamuthia mandrillaris and anti-Naegleria fowleri effects of drugs conjugated with various nanostructures.

Balamuthia mandrillaris and Naegleria fowleri are protist pathogens that can cause fatal infections. Despite mortality rate of > 90%, there is no effective therapy. Treatment remains problematic involving repurposed drugs, e.g., azoles, amphotericin B and miltefosine but requires early diagnosis. In addition to drug discovery, modifying existing drugs using nanotechnology offers promise in the development of therapeutic interventions against these parasitic infections. Herein, various drugs conjugated with nanoparticles were developed and evaluated for their antiprotozoal activities. Characterizations of the drugs' formulations were accomplished utilizing Fourier-transform infrared spectroscopy, efficiency of drug entrapment, polydispersity index, zeta potential, size, and surface morphology. The nanoconjugates were tested against human cells to determine their toxicity in vitro. Themajority of drug nanoconjugates exhibited amoebicidal effects against B. mandrillaris and N. fowleri. Amphotericin B-, Sulfamethoxazole-, Metronidazole-based nanoconjugates are of interest since they exhibited significant amoebicidal effects against both parasites (p < 0.05). Furthermore, Sulfamethoxazole and Naproxen significantly diminished host cell death causedby B. mandrillaris by up to 70% (p < 0.05), while Amphotericin B-, Sulfamethoxazole-, Metronidazole-based drug nanoconjugates showed the highest reduction in host cell death caused by N. fowleri by up to 80%. When tested alone, all of the drug nanoconjugates tested in this study showed limited toxic effects against human cells in vitro (less than 20%). Although these are promising findings, prospective work is warranted to comprehend the mechanistic details of nanoconjugates versus amoebae as well as their in vivo testing, to develop antimicrobials against the devastating infections caused by these parasites.

Azole and 5-nitroimidazole based nanoformulations are potential antiamoebic drug candidates against brain eating amoebae.

Herein, the anti-parasitic activity of azoles (fluconazole and itraconazole) and 5-nitroimdazole (metronidazole) against brain eating amoebae: Naegleria fowleri and Balamuthia mandrillaris was elucidated. Azoles and 5-nitroimdazole based nanoformulations were synthesized and characterized using UV-visible spectrophotometer, atomic force microscopy and fourier transform infrared spectroscopy. H1-NMR, EI-MS and ESI-MS were performed to determine their molecular mass and elucidate their structures. Their size, zeta potential, size distribution and polydispersity index were assessed. Amoebicidal assays revealed that all the drugs and their nanoformulations, (except itraconazole) presented significant anti-amoebic effects against B. mandrillaris, while all the treatments indicated notable amoebicidal properties against N. fowleri. Amoebicidal effects were radically enhanced upon conjugating the drugs with nanoparticles. The IC50 values for KM-38-AgNPs-F, KM-20-AgNPs-M and KM-IF were 65.09µg.mL-1, 91.27µg.mL-1 and 72.19µg.mL-1 respectively against B. mandrillaris. Whereas against N. fowleri, IC50 values were: 71.85µg.mL-1, 73.95µg.mL-1, and 63.01µg.mL-1 respectively. Additionally, nanoformulations significantly reduced N. fowleri mediated host cell death, while nanoformulations along with fluconazole and metronidazole considerably reduced Balamuthia mediated human cell damage. Finally, all the tested drugs and their nanoformulations revealed limited cytotoxic activity against human cerebral microvascular endothelial cell (HBEC-5i) cells. These compounds should be developed into novel chemotherapeutic options for use against these distressing infections due to free-living amoebae, as currently there are no effective treatments.

Cinnamic acid and lactobionic acid based nanoformulations as a potential antiamoebic therapeutics

Acanthamoeba castellanii causes granulomatous amoebic encephalitis, an uncommon but severe brain infection and sight-threatening Acanthamoeba keratitis. Most of the currently used anti-amoebic treatments are not always effective, due to persistence of the cyst stage, and recurrence can occur. Here in this study we synthesize cinnamic acid and lactobionic acid-based magnetic nanoparticles (MNPs) using co-precipitation technique. These nanoformulations were characterized by Fourier transform infrared spectroscopy and Atomic form microscopy. The drugs alone (Hesperidin, Curcumin and Amphotericin B), magnetic NPs alone, and drug-loaded nano-formulations were evaluated at a concentration of 100 μg/mL for antiamoebic activity against a clinical isolate of A. castellanii. Amoebicidal assays revealed that drugs and conjugation of drugs and NPs further enhanced amoebicidal effects of drug-loaded nanoformulations. Drugs and drug-loaded nanoformulations inhibited both encystation and excystation of amoebae. In addition, drugs and drug-loaded nanoformulations inhibited parasite binding capability to the host cells. Neither drugs nor drug-loaded nanoformulations showed cytotoxic effects against host cells and considerably reduced parasite-mediated host cell death. Overall, these findings imply that conjugation of medically approved drugs with MNPs produce potent anti-Acanthamoebic effects, which could eventually lead to the development of therapeutic medications.

Phragmites australis (Cav.) Trin. ex Steud. Extract Induces Apoptosis-like Programmed Cell Death in Acanthamoeba castellanii Trophozoites.

Acanthamoeba keratitis (AK) is an infectious ocular disease which is difficult to diagnose correctly and cure. Development of an effective and safe therapeutic drug for AK is needed. Our preliminary screening of more than 200 extracts from wild plants collected in Korea suggested the potential amoebicidal activity of Phragmites australis (Cav.) Trin. ex Steud. extract (PAE) against Acanthamoeba species. Here, we aimed to analyze the amoebicidal activity of PAE on Acanthamoeba and its underlying amoebicidal mechanism. PAE induced amoebicidal activity against both A. castellanii and A. polyphaga trophozoites, while it showed low cytotoxicity in human corneal epithelial cells (HCE-2) and human retinal pigment epithelial cells (ARPE-19). Transmission electron microscopy analysis showed subcellular morphological changes, such as increased granules, abnormal mitochondria, and atypical cyst wall formation, in the PAE-treated A. castellanii. Fluorometric apoptosis assay and TUNEL assay revealed apoptosis-like programmed cell death (PCD) in the PAE-treated A. castellanii. The PAE treatment increased reactive oxygen species production and reduced mitochondrial membrane potential in the amoeba. The enhanced expression of autophagy-associated genes was also detected. These results suggested that PAE exerted a promising amoebicidal effect on A. castellanii trophozoites via the PCD pathway. PAE could be a potential candidate for developing a therapeutic drug for AK.

Antiamoebic Properties of Ceftriaxone and Zinc-Oxide-Cyclodextrin-Conjugated Ceftriaxone.

Acanthamoeba castellanii is a ubiquitous free-living amoeba capable of instigating keratitis and granulomatous amoebic encephalitis in humans. Treatment remains limited and inconsistent. Accordingly, there is a pressing need for novel compounds. Nanotechnology has been gaining attention for enhancing drug delivery and reducing toxicity. Previous work has shown that various antibiotic classes displayed antiamoebic activity. Herein, we employed two antibiotics: ampicillin and ceftriaxone, conjugated with the nanocarrier zinc oxide and β-cyclodextrin, and tested them against A. castellanii via amoebicidal, amoebistatic, encystment, excystment, cytopathogenicity, and cytotoxicity assays at a concentration of 100 μg/mL. Notably, zinc oxide β-cyclodextrin ceftriaxone significantly inhibited A. castellanii growth and cytopathogenicity. Additionally, both zinc oxide β-cyclodextrin ceftriaxone and ceftriaxone markedly inhibited A. castellanii encystment. Furthermore, all the tested compounds displayed negligible cytotoxicity. However, minimal anti-excystment or amoebicidal effects were observed for the compounds. Accordingly, this novel nanoconjugation should be employed in further studies in hope of discovering novel anti-Acanthamoeba compounds.

The Anti-Amoebic Activity of a Peptidomimetic against Acanthamoeba castellanii.

Acanthamoeba is a free-living protozoan known to cause keratitis most commonly, especially among contact lens wearers. Treatment of Acanthamoeba keratitis is challenging as Acanthamoeba can encyst from the active form, a trophozoite, into a hibernating cyst that is refractory to antibiotics and difficult to kill; therefore, there is a need for more effective anti-amoebic strategies. In this study, we have evaluated the anti-amoebic activity of the antimicrobial peptide mimic RK-758 against Acanthamoeba castellanii. RK-758 peptidomimetic was subjected to biological assays to investigate its amoebicidal, amoebistatic, anti-encystation, and anti-excystation effects on A. castellanii. The anti-amoebic activity of the peptide mimic RK-758 was compared with chlorhexidine against the Acanthamoeba castellanii ATCC30868 and Acanthamoeba castellanii 044 (a clinical strain) with the concentrations of both ranging from 125 µM down to 7.81 µM. All experiments were performed in duplicate with three independent replicates. The data were represented as mean ± SE and analysed using a two-sample t-test and two-tailed distributions. A p < 0.05 was considered statistically significant. The peptidomimetic RK-758 had anti-Acanthamoeba activity against both trophozoites and cysts in a dose-dependent manner. The RK-758 had amoebicidal and growth inhibitory activities of ≥50% at a concentration between 125 µM and 15.6 µM against the trophozoites of both Acanthamoeba strains. Inhibitory effects on the cyst formation and trophozoite re-emergence from cysts were noted at similar concentrations. Chlorhexidine had 50% activity at 7.81 µM and above against the trophozoites and cysts of both strains. In the haemolysis assay, the RK-758 lysed horse RBCs at concentrations greater than 50 µM whereas lysis occurred at concentrations greater than 125 µM for the chlorhexidine. The peptidomimetic RK-758, therefore, has activity against both the trophozoite and cyst forms of Acanthamoeba and has the potential to be further developed as an anti-microbial agent against Acanthamoeba. RK-758 may also have use as an anti-amoebic disinfectant in contact lens solutions.

Imidazothiazole Derivatives Exhibited Potent Effects against Brain-Eating Amoebae

Naegleria fowleri (N. fowleri) is a free-living, unicellular, opportunistic protist responsible for the fatal central nervous system infection, primary amoebic meningoencephalitis (PAM). Given the increase in temperatures due to global warming and climate change, it is estimated that the cases of PAM are on the rise. However, there is a current lack of awareness and effective drugs, meaning there is an urgent need to develop new therapeutic drugs. In this study, the target compounds were synthesized and tested for their anti-amoebic properties against N. fowleri. Most compounds exhibited significant amoebicidal effects against N. fowleri; for example, 1h, 1j, and 1q reduced N. fowleri's viability to 15.14%, 17.45% and 28.78%, respectively. Furthermore, the majority of the compounds showed reductions in amoeba-mediated host death. Of interest are the compounds 1f, 1k, and 1v, as they were capable of reducing the amoeba-mediated host cell death to 52.3%, 51%, and 56.9% from 100%, respectively. Additionally, these compounds exhibit amoebicidal properties as well; they were found to decrease N. fowleri's viability to 26.41%, 27.39%, and 24.13% from 100%, respectively. Moreover, the MIC50 values for 1e, 1f, and 1h were determined to be 48.45 µM, 60.87 µM, and 50.96 µM, respectively. Additionally, the majority of compounds were found to exhibit limited cytotoxicity, except for 1l, 1o, 1p, 1m, 1c, 1b, 1zb, 1z, 1y, and 1x, which exhibited negligible toxicity. It is anticipated that these compounds may be developed further as effective treatments against these devastating infections due to brain-eating amoebae.

Antiamoebic Activity of Imidazothiazole Derivatives against Opportunistic Pathogen Acanthamoeba castellanii.

We examined the antiamoebic effect of several imidazothiazole derivatives on Acanthamoeba castellanii of the T4 genotype. Trypan blue exclusion assays and haemocytometer counting were used to determine the reduction in A. castellanii trophozoite proliferation, in response to treatment with these compounds. To determine the effects of these compounds on host cells, lactate dehydrogenase assay was performed using HeLa cell lines. Amoebicidal assays revealed that the tested compounds at concentrations of 50 µM significantly inhibited amoebae trophozoites compared to controls. Compounds 1m and 1zb showed the highest amoebicidal effects eradicating 70% and 67% of A. castellanii, respectively. The compounds blocked both the encystation and excystation process in A. castellanii. Compounds 1m and 1zb blocked 61% and 55%, respectively, of amoeba binding to human cells. Moreover, the compounds showed minimal cytotoxic effects against host cells and considerably reduced amoeba-mediated host cell death. Overall, our study revealed that compounds 1m and 1zb have excellent antiamoebic potential, and should be considered in the development of curative antiamoebic medications in future studies. Further work is critical to determine the translational value of these findings.

Antiamoebic properties of salicylic acid-based deep eutectic solvents for the development of contact lens disinfecting solutions against Acanthamoeba

Acanthamoeba castellanii is a protist pathogen that can cause sight-threatening keratitis and a fatal infection of the central nervous system, known as granulomatous amoebic encephalitis. In this study, effects of five malonic acid and salicylic acid-based deep eutectic solvents (DES) on A. castellanii were investigated. These are salicylic acid-trioctylphosphine (DES 1), salicylic acid- trihexylamine (DES 2), salicylic acid-trioctylamine (DES 3), malonic acid-trioctylphosphine (DES 4) and malonic acid-trihexylamine (DES 5). The experiments were done by performing amoebicidal, encystment, excystment, cytopathogenicity, and cytotoxicity assays. At micromolar dosage, the solvents DES 2 and DES 3 displayed significant amoebicidal effects (P < 0.05), inhibited encystment and excystment, undermined the cell-mediated cytopathogenicity of A. castellanii, and also displayed minimal cytotoxicity to human cells. Conversely, the chemical components of these solvents: salicylic acid, trihexylamine, and trioctylamine showed minimal effects when tested individually. These results are very promising and to the best of our knowledge, are reported for the first time on the effects of deep eutectic solvents on amoebae. These results can be applied in the development of new formulations of novel contact lens disinfectants against Acanthamoeba castellanii.