Tooba Mahboob

Acanthamoeba spp., free‐living amoebae (FLA), are the causative agents of Acanthamoeba keratitis (AK) and granulomatous amoebic encephalitis (GAE). These diseases have limited treatment options; therefore, drug discovery remains a critical priority. This study evaluated the leaf extract of Annona muricata L., which contains quercetin, formulated in silica nanoparticles against Acanthamoeba species. The anti‐ Acanthamoeba effect of the nanoformulation was determined by its MIC and percentage viability. Scanning electron microscopy confirmed the nanocompound‐directed effect on trophozoites and cysts. The nanoquercetin exhibited anti‐ Acanthamoeba activity, showing greater effectiveness against the trophozoites of Acanthamoeba polyphaga ATCC 30461 (AP) (MIC > 2 mg/mL) compared to cysts (MIC > 8 mg/mL) and against A. triangularis WU19001 (AT) trophozoites (MIC > 4 mg/mL) compared to cysts (MIC > 8 mg/mL). It also showed reduced activity against Acanthamoeba castellanii ATCC 30010 (AC3) trophozoites (MIC > 4 mg/mL) compared to cysts (MIC > 8 mg/mL) and A. castellanii ATCC50739 (AC5) trophozoites (MIC > 4 mg/mL) compared to cysts (MIC > 8 mg/mL). The anti‐ Acanthamoeba activity of A. muricata on all four species was compared to chlorhexidine, which had MIC values of 0.008 mg/mL for AC3 and AT trophozoites and 0.016 mg/mL for AC5 and AP trophozoites. For cysts, chlorhexidine exhibited MIC values of > 0.0128 mg/mL for AC3 and AP, 0.032 mg/mL for AC5, and 0.064 mg/mL for AT. All MIC values demonstrated inhibition of Acanthamoeba species with reduced viability exceeding 90%. SEM findings revealed morphological changes in Acanthamoeba , indicating parasitic inhibition. Quercetin‐loaded silica nanoparticles demonstrated anti‐ Acanthamoeba activity with low cytotoxicity. This indicates the potential of quercetin as a nano‐enhanced natural therapeutic or contact lens‐care agent. This study is limited by its in vitro design, small parasite strain panel, and potential nanoparticle variability. This highlights the need for in vivo validation, ocular/contact lens model testing, and formulation optimization for improved cysticidal efficacy.

Background and Aim: Malaria continues to pose a global health challenge, exacerbated by the emergence of drug-resistant strains of Plasmodium falciparum. This study aimed to evaluate the anti-Plasmodium potential of Propolis extracts collected from various Iranian regions and to characterize the molecular interactions of their bioactive phytochemicals with P. falciparum lactate dehydrogenase (PfLDH), a key enzyme in parasite glycolysis. Materials and Methods: The anti-Plasmodium activity of ethanol-extracted Propolis was assessed against P. falciparum NF54 using the SYBR Green I fluorescence assay. Gas chromatography-mass spectrometry (GC-MS) analysis identified major phytochemicals in the most active extract. Molecular docking and 100-ns molecular dynamic (MD) simulations were performed to evaluate the binding affinity and stability of selected compounds (tectochrysin and galangin) against PfLDH in both holo (Protein Data Bank [PDB] ID: 1LDG) and apo (PDB ID: 2X8L) forms. Results: Propolis collected from Kermanshah city exhibited the highest anti-Plasmodium activity (IC50 = 6.69 ± 1.44 μg/mL). GC-MS analysis identified tectochrysin and galangin as major constituents. Molecular docking revealed strong binding affinities of tectochrysin (−7.8 kcal/mol) and galangin (−7.5 kcal/mol) to PfLDH, surpassing the binding energies of standard antimalarial drugs (chloroquine and quinine). MD simulations confirmed the stability of tectochrysin and galangin within the PfLDH active sites, with favorable root mean square deviation, root mean square fluctuation, gyration, solvent-accessible surface area, molecular surface area, and polar surface area profiles, indicating persistent and stable protein-ligand interactions throughout the simulation. Conclusion: The findings support the promising anti-Plasmodium potential of Propolis-derived compounds, particularly tectochrysin and galangin, as novel PfLDH inhibitors. Their potential applicability in transdisciplinary anti-parasitic therapy across human and veterinary medicine warrants further in vivo validation and clinical investigations. Keywords: anti-malarial candidates, galangin, lactate dehydrogenase, molecular docking, molecular dynamics, Plasmodium falciparum, Propolis extract, tectochrysin

Medicinal plants have been used as alternative agents for the treatment of infections. This study aimed to investigate bioactivities of medicinal plant extracts including Knema retusa extract (Kre) against Acanthamoeba triangularis T4 in vitro and in silico . Anti‐ Acanthamoeba activities of 44 extracts from 5 plant species were determined. From 44 tested extracts, a chloroform extract of Kre bark showed the strongest anti‐ Acanthamoeba activities against both trophozoites and cysts, with MIC values of 32.25 and 62.50 μg/mL, respectively. Then, amebicidal and antiadhesion activities of Kre against A. triangularis were investigated. Kre reduced the growth by 3 logs within 8 h at 4 × MIC. Disruption of the cells with abnormal shapes was observed when trophozoites were treated with Kre. Trophozoites had lost their robust acanthopodia and began to shrink after treatment with Kre. Treated cysts exhibited wall disruption and dramatically showed forms of marked retraction. Treatment of Kre at 1/2 × MIC showed about 87% reduction in the trophozoite adhesion, while treatment at 2 × MIC exhibited a 59% reduction in the trophozoite adhesion to the plastic surface, compared with the control. Furthermore, 1 log cells/mL (90%) of the contact lens adhesive trophozoites were reduced and removed after treatment with Kre. Molecular docking indicated that E2N, the main compound in Kre, exhibited strong binding to the ligand binding sites at β‐tubulin, with a binding energy of −7.01 kcal/mol and an inhibitory constant of 2.43–7.32 μM. E2N generated multiple connections via hydrogen, hydrophobic, ionic, and water bridge bonding and maintained these connections until the simulation finished, facilitating the creation of stable bindings with the β‐tubulin protein as measured by molecular dynamics simulation. These findings suggest that Kre exhibits amebicidal and antiadhesion activities which could be used for the prevention of A. triangularis adhesion to contact lenses.

Noncanonical autophagy including unconventional protein secretion has been extensively studied. Our work focused on a leaderless IL‐1β protein secretion from human macrophage in response to Acanthamoeba castellanii components, Acanthamoeba culture supernatant (CS) and cell lysate (CL), as well as its association with macrophage autophagy. Phorbol 12‐myristate 13‐acetate (PMA)–induced THP‐1 macrophages were treated with Acanthamoeba components of pathogenic (ATCC50739) and nonpathogenic (ATCC30010) strains in vitro. The data showed that Acanthamoeba treatment resulted in low IL‐1β secretion from macrophages. In addition, Acanthamoeba CL of both strains was able to upregulate autophagy‐related (Atg) protein 8, an autophagy marker, whereas Acanthamoeba CS downregulated Atg8 expression. We further manipulated autophagy and found that autophagy induction by starvation diminished IL‐1β secretion while autophagy inhibition by 3‐methyladenine (3MA) increased IL‐1β secretion. Interestingly, in the presence of Acanthamoeba components either under starvation or 3MA treatment, IL‐1β secretion was significantly reduced. Transcriptional expression of other ATG genes, i.e., ATG6, ATG7, and ATG5, were investigated and showed that their mRNA expression was maintained at the basal level under A. castellanii CS or CL treatment. Inflammasome‐related genes, NLRP3 and CASPASE1, were upregulated following A. castellanii 50739 CS treatment but not in A. castellanii 50739 CL‐treated condition. However, both conditions were able to increase IL‐1β mRNA expression. TEM micrographs revealed that 3MA treatment induced the formation of large vacuoles and accumulation of autophagosome at the edge of THP‐1 macrophages. However, the number and size of their structures were declined in the presence of A. castellanii 50739 CS with 3MA. Furthermore, immunofluorescence staining demonstrated the association between Atg8/LC3 and IL‐1β expression, where downregulation of Atg8 by A. castellanii 50739 CS led to the upregulation of IL‐1β. Altogether, the data indicate that Acanthamoeba can manipulate macrophage autophagy, thereby controlling low IL‐1β secretion. The expression of autophagy‐ and inflammasome‐related genes also indicates multiple mechanisms in IL‐1β secretion in response to Acanthamoeba components. However, further characterization of Atg proteins and investigations into other intracellular pathways or defense mechanisms are needed to fully understand the unconventional secretion of IL‐1β in macrophages. This knowledge could eventually lead to the development of innovative therapeutic strategies against Acanthamoeba infection by modulating autophagy or macrophage responses.

Background and Aim: Keratitis is a serious ocular infection often caused by pathogenic micro-organisms such as Acanthamoeba spp. Among other harmful microbes, Acanthamoeba keratitis presents a particular challenge due to its resistance to conventional antimicrobial agents. Piper betle Linn., commonly known as betel leaf, has been traditionally used for its medicinal properties. This study aimed to assess the potential of the leaf ethanol extract of P. betle Linn. in the treatment of Acanthamoeba triangularis in monoculture and co-culture with two prevalent pathogenic bacteria, Staphylococcus aureus and Pseudomonas aeruginosa, associated with keratitis. Materials and Methods: Minimum inhibitory concentrations (MICs) of A. triangularis, S. aureus, and P. aeruginosa extracts in monoculture and coinfected conditions were examined. In addition, this study explored the potential of the extract in preventing Acanthamoeba adherence in both monoculture and co-culture environments. Scanning electron microscopy (SEM) analysis confirmed the impact of the extract on Acanthamoeba cell membranes, including acanthopodia. Furthermore, a time-kill kinetic assay was used to validate the amoebicidal activity of the extract against A. triangularis and the tested bacteria. Results: MICs for trophozoites, cysts, P. aeruginosa, and S. aureus in the monoculture were 0.25, 0.25, 0.51, and 0.128 mg/mL, respectively, whereas the MICs for Acanthamoeba coinfected with bacteria were higher than those in the monoculture. This extract inhibited the growth of A. triangularis trophozoites and cysts for up to 72 h. Moreover, P. betle extract effectively prevented the adherence of Acanthamoeba to contact lenses under monoculture conditions. SEM analysis confirmed that P. betle extract affects the cell membrane of Acanthamoeba, including Acanthopodia. In addition, the time-kill kinetic assay confirmed that the extract contained amoebicidal activity against A. triangularis, including the tested bacteria. Notably, S. aureus was more susceptible than A. triangularis and P. aeruginosa to P. betle extract treatment. Unexpectedly, our study revealed that S. aureus negatively affected A. triangularis in the co-culture after 3 days of incubation, whereas P. aeruginosa facilitated the growth of A. triangularis in the presence of the extract. Conclusion: This study provides compelling evidence of the anti-adhesive and anti-Acanthamoeba properties of P. betle leaf extract against A. triangularis under monoculture and co-culture conditions. The observed impact on Acanthamoeba cell membranes, coupled with the time-kill kinetic assay results, underscores the potential of P. betle leaf extract as a promising agent for combating Acanthamoeba-related infections in humans and animals. Keywords: Piper betle extract, Acanthamoeba triangularis, co-infection, keratitis, Pseudomonas aeruginosa, Staphylococcus aureus.

The WHO declared coronavirus disease 2019 (COVID-19) a pandemic in March 2020, which was caused by novel coronavirus severe acute respiratory coronavirus 2 (SARS-CoV-2). SARS-CoV-2 made its first entry into the world in November 2019, and the first case was detected in Wuhan, China. Mutations in the SARS-CoV-2 genome distressed life in almost every discipline by the extended production of novel viral variants. In this article, authorized SARS-CoV-2 vaccines including mRNA vaccines, DNA vaccines, subunit vaccines, inactivated virus vaccines, viral vector vaccine, live attenuated virus vaccines and mix and match vaccines will be discussed based on their mechanism, administration, storage, stability, safety and efficacy. The information was collected from various journals via electronic searches including PubMed, Science Direct, Google Scholar and the WHO platform. This review article includes a brief summary on the pathophysiology, epidemiology, mutant variants and management strategies related to COVID-19. Due to the continuous production and unsatisfactory understanding of novel variants of SARS-CoV-2, it is important to design an effective vaccine along with long-lasting protection against variant strains by eliminating the gaps through practical and theoretical knowledge. Consequently, it is mandatory to update the literature through previous and ongoing trials of vaccines tested among various ethnicities and age groups to gain a better insight into management strategies and combat complications associated with upcoming novel variants of SARS-CoV-2.

Background and Objectives: Ochradenus baccatus belongs to the family Resedaceae. It is widely spread in Saudi Arabia and other countries in Southwest Asia. O. baccatus is extensively used in traditional medicine as an anti-inflammatory and antibacterial agent, in addition to being a vital source of food for certain desert animal species. The aim of the present study was to investigate the chemical composition and antibacterial/anticancer activities of O. baccatus methanolic extracts collected from Hail, Saudi Arabia. Materials and Methods: The O. baccatus extracts were obtained by macerating the crude powder in methanol, followed by filtration and evaporation. Liquid chromatography–mass spectrometry (LC-MS) was used to analyze the methanolic extracts’ chemical constituents. Broth microdilution assay for minimum inhibitory concentration (MIC) determination was used to assess antimicrobial activity, while the extracts’ anticancer potential was assessed by sulforhodamine B Assay (SRB) assay. Results: The results of the antibacterial assay showed that the methanolic extracts from the roots and branches possessed varying degrees of activity against particular bacterial strains, with the highest activity being exerted by the branches’ extract against Escherichia coli and Salmonella typhimurium (St), demonstrating MIC values of 15.6 µg/mL and 20 µg/mL, respectively. Furthermore, the SRB cell viability assay revealed that only the branches’ extract inhibited the growth of A549 cancer cells, with an IC₅₀ value of 86.19 µg/mL. The LC-MS analysis of the methanolic extracts from the plant’s roots and branches was then conducted, resulting in the identification of 8 and 13 major chemical constituents, respectively. Azelaic acid, β-amyrin, and phytanic acid are some of the bioactive compounds that were detected in the extracts via LC-MS, and they are thought to be responsible for the observed antibacterial/anticancer activity of O. baccatus methanolic extracts. Conclusions: This study confirmed the antibacterial/anticancer potential of O. baccatus methanolic extracts and analyzed their phytochemical constituents. Further isolation and biological screening are warranted to understand the therapeutic potential of O. baccatus.

Among the large number of plants that are part of the Ayurvedic system of medicine in India and Bangladesh, Zingiber officinale Roscoe (Zingiberaceae), or ginger in English, holds a special place and is often referred to as “Mahaushadha” (great medicine) and “Vishvabhesaja” (worldwide or universal herb) to signify its special status. The plant and particularly its rhizomes are used both in the raw and dry form for the relief of a multitude of disorders. Since a number of these disorders occur in patients infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), it was of interest to perform in silico studies (molecular docking) to evaluate the binding affinities of a number of constituents of Zingiber officinale with the 3C-like protease or main protease (Mpro) of SARS-CoV-2, which plays an essential role in the cleavage of viral polyproteins and subsequent viral replication. Our studies indicated that 2 of the compounds present in ginger, namely, chlorogenic acid and hesperidin, had high binding affinities for Mpro with predicted binding energies of −7.5 and −8.3 kcal/mol. The two-dimensional and three-dimensional interactions also showed that, while chlorogenic acid interacts with one of the His41 amino acids of the catalytic dyad of Mpro, hesperidin interacts with the other amino acid Cys145, which can account for their predicted high binding energies and, therefore, possibly can inhibit Mpro activity. Taken together, our findings indicate that ginger, besides alleviating the symptoms induced by SARS-CoV-2, may also play a role in inhibiting the virus.