Tracking Five Millennia of Horse Management with Extensive Ancient Genome Time Series.

Horse domestication revolutionized warfare and accelerated travel, trade, and the geographic expansion of languages. Here, we present the largest DNA time series for a non-human organism to date, including genome-scale data from 149 ancient animals and 129 ancient genomes (≥1-fold coverage), 87 of which are new. This extensive dataset allows us to assess the modern legacy of past equestrian civilizations. We find that two extinct horse lineages existed during early domestication, one at the far western (Iberia) and the other at the far eastern range (Siberia) of Eurasia. None of these contributed significantly to modern diversity. We show that the influence of Persian-related horse lineages increased following the Islamic conquests in Europe and Asia. Multiple alleles associated with elite-racing, including at the MSTN "speed gene," only rose in popularity within the last millennium. Finally, the development of modern breeding impacted genetic diversity more dramatically than the previous millennia of human management.

Widespread horse-based mobility arose around 2,200 BCE in Eurasia.

Horses revolutionized human history with fast mobility1. However, the timeline between their domestication and widespread integration as a means of transportation remains contentious2-4. Here we assemble a large collection of 475 ancient horse genomes to assess the period when these animals were first reshaped by human agency in Eurasia. We find that reproductive control of the modern domestic lineage emerged ~2,200 BCE (Before Common Era), through close kin mating and shortened generation times. Reproductive control emerged following a severe domestication bottleneck starting no earlier than ~2,700 BCE, and coincided with a sudden expansion across Eurasia that ultimately resulted in the replacement of nearly every local horse lineage. This expansion marked the rise of widespread horse-based mobility in human history, which refutes the commonly-held narrative of large horse herds accompanying the massive migration of steppe peoples across Europe ~3,000 BCE and earlier3,5. Finally, we detect significantly shortened generation times at Botai ~3,500 BCE, a settlement from Central Asia associated with corrals and a subsistence economy centered on horses6,7. This supports local horse husbandry before the rise of modern domestic bloodlines.

The origins and spread of domestic horses from the Western Eurasian steppes.

Domestication of horses fundamentally transformed long-range mobility and warfare1. However, modern domesticated breeds do not descend from the earliest domestic horse lineage associated with archaeological evidence of bridling, milking and corralling2-4 at Botai, Central Asia around 3500 BC3. Other longstanding candidate regions for horse domestication, such as Iberia5 and Anatolia6, have also recently been challenged. Thus, the genetic, geographic and temporal origins of modern domestic horses have remained unknown. Here we pinpoint the Western Eurasian steppes, especially the lower Volga-Don region, as the homeland of modern domestic horses. Furthermore, we map the population changes accompanying domestication from 273 ancient horse genomes. This reveals that modern domestic horses ultimately replaced almost all other local populations as they expanded rapidly across Eurasia from about 2000 BC, synchronously with equestrian material culture, including Sintashta spoke-wheeled chariots. We find that equestrianism involved strong selection for critical locomotor and behavioural adaptations at the GSDMC and ZFPM1 genes. Our results reject the commonly held association7 between horseback riding and the massive expansion of Yamnaya steppe pastoralists into Europe around 3000 BC8,9 driving the spread of Indo-European languages10. This contrasts with the scenario in Asia where Indo-Iranian languages, chariots and horses spread together, following the early second millennium BC Sintashta culture11,12.

Observation of Period-Doubling Bloch Oscillations.

Bloch oscillations refer to the periodic oscillation of a wave packet in a lattice under a constant force. Typically, the oscillation has a fundamental period that corresponds to the wave packet traversing the first Brillouin zone once. Here, we demonstrate, both theoretically and experimentally, the optical Bloch oscillations where the wave packet must traverse the first Brillouin zone twice to complete a full cycle, resulting in a period of oscillation that is 2 times longer than that of usual Bloch oscillations. The unusual Bloch oscillations arise due to the band crossing of valley-Hall topological edge states at the Brillouin boundary for zigzag domain walls between two staggered honeycomb lattices with inverted on-site energy detuning, which are protected by the glide-reflection symmetry of the underlying structures. Our work sheds light on the direct detection of band crossings resulting from intrinsic symmetries that extend beyond the fundamental translational symmetry in topological systems.

Self-assembled micelles loaded with itraconazole as anti-Acanthamoeba nano-formulation.

Acanthamoeba castellanii are opportunistic pathogens known to cause infection of the central nervous system termed: granulomatous amoebic encephalitis, that mostly effects immunocompromised individuals, and a sight threatening keratitis, known as Acanthamoeba keratitis, which mostly affects contact lens wearers. The current treatment available is problematic, and is toxic. Herein, an amphiphilic star polymer with AB2 miktoarms [A = hydrophobic poly(ℇ-Caprolacton) and B = hydrophilic poly (ethylene glycol)] was synthesized by ring opening polymerization and CuI catalyzed azide-alkyne cycloaddition. Characterization by 1H and 13C NMR spectroscopy, size-exclusion chromatography and fluorescence spectroscopy was accomplished. The hydrophobic drug itraconazole (ITZ) was incorporated in self-assembled micellar structure of AB2 miktoarms through co-solvent evaporation. The properties of ITZ loaded (ITZ-PCL-PEG2) and blank micelles (PCL-PEG2) were investigated through zeta sizer, scanning electron microscopy and Fourier-transform infrared spectroscopy. Itraconazole alone (ITZ), polymer (DPB-PCL), empty polymeric micelles (PCL-PEG2) alone, and itraconazole loaded in polymeric micelles (ITZ-PCL-PEG2) were tested for anti-amoebic potential against Acanthamoeba, and the cytotoxicity on human cells were determined. The polymer was able to self-assemble in aqueous conditions and exhibited low value for critical micelle concentration (CMC) 0.05-0.06 µg/mL. The maximum entrapment efficiency of ITZ was 68%. Of note, ITZ, DPB, PCL-PEG2 and ITZ-PCL-PEG2 inhibited amoebae trophozoites by 37.34%, 36.30%, 35.77%, and 68.24%, respectively, as compared to controls. Moreover, ITZ-PCL-PEG2 revealed limited cytotoxicity against human keratinocyte cells. These results are indicative that ITZ-PCL-PEG2 micelle show significantly better anti-amoebic effects as compared to ITZ alone and thus should be investigated further in vivo to determine its clinical potential.

Antibacterial effects of quercetagetin are significantly enhanced upon conjugation with chitosan engineered copper oxide nanoparticles.

The development of antibiotic alternatives that entail distinctive chemistry and modes of action is necessary due to the threat posed by drug resistance. Nanotechnology has gained increasing attention in recent years, as a vehicle to enhance the efficacy of existing antimicrobials. In this study, Chitosan copper oxide nanoparticles (CHI-CuO) were synthesized and were further loaded with Quercetagetin (QTG) to achieve the desired (CHI-CuO-QTG). Size distribution, zeta potential and morphological analysis were accomplished. Next, the developed CHI-CuO-QTG was assessed for synergistic antibacterial properties, as well as cytotoxic attributes. Bactericidal assays revealed that CHI-CuO conjugation showed remarkable effects and enhanced QTG effects against a range of Gram + ve and Gram - ve bacteria. The MIC50 of QTG against S. pyogenes was 107 µg/mL while CHI-CuO-QTG reduced it to 9 µg/mL. Similar results were observed when tested against S. pneumoniae. Likewise, the MIC50 of QTG against S. enterica was 38 µg/mL while CHI-CuO-QTG reduced it to 7 µg/mL. For E. coli K1, the MIC50 of QTG was 42 µg/mL while with CHI-CuO-QTG it was 23 µg/mL. Finally, the MIC50 of QTG against S. marcescens was 98 µg/mL while CHI-CuO-QTG reduced it to 10 µg/mL. Notably, the CHI-CuO-QTG nano-formulation showed limited damage when tested against human cells using lactate dehydrogenase release assays. Importantly, bacterial-mediated human cell damage was reduced by prior treatment of bacteria using drug nano-formulations. These findings are remarkable and clearly demonstrate that drug-nanoparticle formulations using nanotechnology is an important avenue in developing potential therapeutic interventions against microbial infections.

Nanomedicine: Patuletin-conjugated with zinc oxide exhibit potent effects against Gram-negative and Gram-positive bacterial pathogens.

With the emergence of drug-resistance, there is a need for novel anti-bacterials or to enhance the efficacy of existing drugs. In this study, Patuletin (PA), a flavanoid was loaded onto Gallic acid modified Zinc oxide nanoparticles (PA-GA-ZnO), and evaluated for antibacterial properties against Gram-positive (Bacillus cereus and Streptococcus pneumoniae) and Gram-negative (Samonella enterica and Escherichia coli) bacteria. Characterization of PA, GA-ZnO and PA-GA-ZnO' nanoparticles was accomplished utilizing fourier-transform infrared spectroscopy, efficiency of drug entrapment, polydispersity index, zeta potential, size, and surface morphology analysis through atomic force microscopy. Using bactericidal assays, the results revealed that ZnO conjugation displayed remarkable effects and enhanced Patuletin's effects against both Gram-positive and Gram-negative bacteria, with the minimum inhibitory concentration observed at micromolar concentrations. Cytopathogenicity assays exhibited that the drug-nanoconjugates reduced bacterial-mediated human cell death with minimal side effects to human cells. When tested alone, drug-nanoconjugates tested in this study showed limited toxic effects against human cells in vitro. These are promising findings, but future work is needed to understand the molecular mechanisms of effects of drug-nanoconjugates against bacterial pathogens, in addition to in vivo testing to determine their translational value. This study suggests that Patuletin-loaded nano-formulation (PA-GA-ZnO) may be implicated in a multi-target mechanism that affects both Gram-positive and Gram-negative pathogen cell structures, however this needs to be ascertained in future work.

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.

Next generation imidazothiazole and imidazooxazole derivatives as potential drugs against brain-eating amoebae.

Managing primary amoebic meningoencephalitis, induced by Naegleria fowleri poses a complex medical challenge. There is currently no specific anti-amoebic drug that has proven effectiveness against N. fowleri infection. Ongoing research endeavours are dedicated to uncovering innovative treatment strategies, including the utilization of drugs and immune modulators targeting Naegleria infection. In this study, we explored the potential of imidazo[2,1-b]thiazole and imidazooxazole derivatives that incorporate sulfonate and sulfamate groups as agents with anti-amoebic properties against N. fowleri. We assessed several synthesized compounds (1f, 1m, 1q, 1s, and 1t) for their efficacy in eliminating amoebae, their impact on cytotoxicity, and their influence on the damage caused to human cerebral microvascular endothelial (HBEC-5i) cells when exposed to the N. fowleri (ATCC 30174) strain. The outcomes revealed that, among the five compounds under examination, 1m, 1q, and 1t demonstrated notable anti-parasitic effects against N. fowleri (P ≤ 0.05). Compound 1t exhibited the highest anti-parasitic activity, reducing N. fowleri population by 80%. Additionally, three compounds, 1m, 1q, and 1t, significantly mitigated the damage inflicted on host cells by N. fowleri. However, the results of cytotoxicity analysis indicated that while 1m and 1q had minimal cytotoxic effects on endothelial cells, compound 1t caused moderate cytotoxicity (34%). Consequently, we conclude that imidazo[2,1-b]thiazole and imidazooxazole derivatives containing sulfonate and sulfamate groups exhibit a marked capacity to eliminate amoebae viability while causing limited toxicity to human cells. In aggregate, these findings hold promise that could potentially evolve into novel therapeutic options for treating N. fowleri infection.

Uncovering Potentially Therapeutic Phytochemicals, In silico Analysis, and Biological Assessment of South-Chinese Red Dragon Fruit (Hylocereus polyrhizus).

Red dragon fruit is gaining popularity globally due to its nutritional value and bioactive components. The study aimed to assess the phytochemical, nutritional composition, antioxidant, antibacterial, and cytotoxic properties of extracts from the South Chinese red dragon fruit peel, flesh, and seeds. Extract fractions with increasing polarity (ethyl acetate

Applications of photodynamic therapy in keratitis.

Keratitis is corneal inflammatory disease which may be caused by several reason such as an injury, allergy, as well as a microbial infection. Besides these, overexposure to ultraviolet light and unhygienic practice of contact lenses are also associated with keratitis. Based on the cause of keratitis, different lines of treatments are recommended. Photodynamic therapy is a promising approach that utilizes light activated compounds to instigate either killing or healing mechanism to treat various diseases including both communicable and non-communicable diseases. This review focuses on clinically-important patent applications and the recent literature for the use of photodynamic therapy against keratitis.

Gut microbiome of Crocodylus porosus and cellular stress: inhibition of nitric oxide, interleukin 1-beta, tumor necrosis factor-alpha, and prostaglandin E2 in cerebrovascular endothelial cells.

Crocodiles are renowned for their resilience and capacity to withstand environmental stressors, likely influenced by their unique gut microbiome. In this study, we determined whether selected gut bacteria of Crocodylus porosus exhibit anti-inflammatory effects in response to stress, by measuring nitric oxide release, interleukin 1-beta, tumor necrosis factor-alpha, and prostaglandin E2 in cerebrovascular endothelial cells. Using the Griess assay, the findings revealed that among several C. porosus gut bacterial isolates, the conditioned media containing the metabolites of two bacterial strains (CP27 and CP36) inhibited nitric oxide production significantly, in response to the positive control, i.e., taxol-treatment. Notably, CP27 and CP36 were more potent at reducing nitric oxide production than senloytic compounds (fisetin, quercetin). Using enzyme linked immunosorbent assays, the production of pro-inflammatory cytokines (IL-1ß, TNF-α, PGE2), was markedly reduced by treatment with CP27 and CP36, in response to stress. Both CP27 and CP36 contain a plethora of metabolites to exact their effects [(3,4-dihydroxyphenylglycol, 5-methoxytryptophan, nifedipine, 4-chlorotestosterone-17-acetate, 3-phenoxypropionic acid, lactic acid, f-Honaucin A, l,l-Cyclo(leucylprolyl), 3-hydroxy-decanoic acid etc.], indicative of their potential in providing protection against cellular stress. Further high-throughput bioassay-guided testing of gut microbial metabolites from crocodiles, individually as well as in combination, together with the underlying molecular mechanisms, in vitro and in vivo will elucidate their value in the rational development of innovative therapies against cellular stress/gut dysbiosis.

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. The majority 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 caused by 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.

Effect of embelin on inhibition of cell growth and induction of apoptosis in Acanthamoeba castellanii.

Acanthamoeba castellanii is the causative agent of fatal encephalitis and blinding keratitis. Current therapies remain a challenge, hence there is a need to search for new therapeutics. Here, we tested embelin (EMB) and silver nanoparticles doped with embelin (EMB-AgNPs) against A. castellanii. Using amoebicidal assays, the results revealed that both compounds inhibited the viability of Acanthamoeba, having an IC50 of 27.16 ± 0.63 and 13.63 ± 1.08 µM, respectively, while causing minimal cytotoxicity against HaCaT cells in vitro. The findings suggest that both samples induced apoptosis through the mitochondria-mediated pathway. Differentially expressed genes analysis showed that 652 genes were uniquely expressed in treated versus untreated cells, out of which 191 were significantly regulated in the negative control vs. conjugate. Combining the analysis, seven genes (ARIH1, RAP1, H3, SDR16C5, GST, SRX1, and PFN) were highlighted as the most significant (Log2 (FC) value ± 4) for the molecular mode of action in vitro. The KEGG analysis linked most of the genes to apoptosis, the oxidative stress signaling pathway, cytochrome P450, Rap1, and the oxytocin signaling pathways. In summary, this study provides a thorough framework for developing therapeutic agents against microbial infections using EMB and EMB-AgNPs.

Alpha-Mangostin and its nano-conjugates induced programmed cell death in Acanthamoeba castellanii belonging to the T4 genotype.

Acanthamoeba are free living amoebae that are the causative agent of keratitis and granulomatous amoebic encephalitis. Alpha-Mangostin (AMS) is a significant xanthone; that demonstrates a wide range of biological activities. Here, the anti-amoebic activity of α-Mangostin and its silver nano conjugates (AMS-AgNPs) were evaluated against pathogenic A. castellanii trophozoites and cysts in vitro. Amoebicidal assays showed that both AMS and AMS-AgNPs inhibited the viability of A. castellanii dose-dependently, with an IC50 of 88.5 ± 2.04 and 20.2 ± 2.17 µM, respectively. Both formulations inhibited A. castellanii-mediated human keratinocyte cell cytopathogenicity. Functional assays showed that both samples caused apoptosis through the mitochondrial pathway and reduced mitochondrial membrane potential and ATP production, while increasing reactive oxygen species (ROS) and nicotinamide adenine dinucleotide phosphate (NADPH) cytochrome-c reductase in the cytosol. Whole transcriptome sequencing of A. castellanii showed the expression of 826 genes, with 447 genes being up-regulated and 379 genes being down-regulated post treatment. The Kyoto Encyclopedia of Genes and Genomes analysis showed that the majority of genes were linked to apoptosis, autophagy, RAP1, AGE-RAGE and oxytocin signalling pathways. Seven genes (PTEN, H3, ARIH1, SDR16C5, PFN, glnA GLUL, and SRX1) were identified as the most significant (Log2 (FC) value 4) for molecular mode of action in vitro. Future in vivo studies with AMS and nanoconjugates are needed to realize the clinical potential of this work.

Anti-amoebic activity of a series of benzofuran/benzothiophene derivatives against Acanthamoeba castellanii belonging to the T4 genotype.

AIMS: To determine the anti-amoebic activity of benzofuran/benzothiophene-possessing compounds against Acanthamoeba castellanii of the T4 genotype. METHOD AND RESULTS: A series of benzofuran/benzothiophene-possessing compounds were tested for their anti-amoebic activities, in particular, to block encystation and excystation processes in amoebae. Cytotoxicity of the compounds were evaluated using lactate dehydrogenase (LDH) assays. The amoebicidal assay results revealed significant anti-amoebic effects against A. castellanii. Compounds 1p and 1e showed the highest amoebicidal activity, eliminating 68% and 64% of the amoebae, respectively. These compounds remarkably repressed both the encystation and excystation processes in A. castellanii. Furthermore, the selected compounds presented minimal cytotoxic properties against human cells, as well as considerably abridged amoeba-mediated cytopathogenicity when compared to the amoebae alone. CONCLUSIONS: Our findings show that benzofuran/benzothiophene derivatives depict potent anti-amoebic activities; thus these compounds should be used as promising and novel agents in the rationale development of therapeutic strategies against Acanthamoeba infections.

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

AIM: Herein, the anti-parasitic activity of azoles (fluconazole and itraconazole) and 5-nitroimdazole (metronidazole) against the brain-eating amoebae: Naegleria fowleri and Balamuthia mandrillaris was elucidated. METHODS AND RESULTS: Azoles and 5-nitroimidazole based nanoformulations were synthesized and characterized using a 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 (PDI) 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 65.09, 91.27, and 72.19 µg.mL-1, respectively, against B. mandrillaris. Whereas against N. fowleri, the IC50 values were: 71.85, 73.95, and 63.01 µg.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. CONCLUSION: 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.

CDK4/6 inhibition triggers anti-tumour immunity.

Cyclin-dependent kinases 4 and 6 (CDK4/6) are fundamental drivers of the cell cycle and are required for the initiation and progression of various malignancies. Pharmacological inhibitors of CDK4/6 have shown significant activity against several solid tumours. Their primary mechanism of action is thought to be the inhibition of phosphorylation of the retinoblastoma tumour suppressor, inducing G1 cell cycle arrest in tumour cells. Here we use mouse models of breast carcinoma and other solid tumours to show that selective CDK4/6 inhibitors not only induce tumour cell cycle arrest, but also promote anti-tumour immunity. We confirm this phenomenon through transcriptomic analysis of serial biopsies from a clinical trial of CDK4/6 inhibitor treatment for breast cancer. The enhanced anti-tumour immune response has two underpinnings. First, CDK4/6 inhibitors activate tumour cell expression of endogenous retroviral elements, thus increasing intracellular levels of double-stranded RNA. This in turn stimulates production of type III interferons and hence enhances tumour antigen presentation. Second, CDK4/6 inhibitors markedly suppress the proliferation of regulatory T cells. Mechanistically, the effects of CDK4/6 inhibitors both on tumour cells and on regulatory T cells are associated with reduced activity of the E2F target, DNA methyltransferase 1. Ultimately, these events promote cytotoxic T-cell-mediated clearance of tumour cells, which is further enhanced by the addition of immune checkpoint blockade. Our findings indicate that CDK4/6 inhibitors increase tumour immunogenicity and provide a rationale for new combination regimens comprising CDK4/6 inhibitors and immunotherapies as anti-cancer treatment.

Molecular characterisation and potential pathogenicity analysis of Acanthamoeba isolated from recreational lakes in Peninsular Malaysia.

The present study aims to identify the Acanthamoeba genotypes and their pathogenic potential in three recreational lakes in Malaysia. Thirty water samples were collected by purposive sampling between June and July 2022. Physical parameters of water quality were measured in situ while chemical and microbiological analyses were performed in the laboratory. The samples were vacuum filtered through nitrate filter, cultured onto non-nutrient agar and observed microscopically for amoebic growth. DNAs from positive samples were extracted and made to react with polymerase chain reaction using specific primers. Physiological tolerance tests were performed for all Acanthamoeba-positive samples. The presence of Acanthamoeba was found in 26 of 30 water samples by PCR. The highest rate in lake waters contaminated with amoeba was in Biru Lake (100%), followed by Titiwangsa Lake (80%) and Shah Alam Lake (80%). ORP, water temperature, pH and DO were found to be significantly correlated with the presence of Acanthamoeba. The most common genotype was T4. Temperature- and osmo-tolerance tests showed that 8 (30.8%) of the genotypes T4, T9 and T11 were highly pathogenic. The presence of genotype T4 in habitats related to human activities supports the relevance of this amoeba as a potential public health concern.

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.