Identification of sheep eimerian parasites, Eimeria crandallis and Eimeria faurei, employing microscopic and molecular tools.

Coccidiosis, caused by apicomplexan Eimeria species, is a protozoan disease that affects various species of wild and domestic animals. However, data available on Eimeria diversity in ruminants in Saudi Arabia is meagre. Therefore, this study was designed to investigate some eimerian parasites infecting sheep (Sawakni and Harrie breeds) using microscopy and molecular methods for the first time in Saudi Arabia. Twenty-four fecal samples were collected from sheep farms. Based on the floatation technique, eimerian oocysts were observed in 8 of the 24 (33.33%) fecal samples. The coccidian-positive samples were subjected to fecal culture in a shallow layer of 2.5% potassium dichromate (K2 Cr2 O7 ). Detected eimerian oocysts were described micromorphometrically as the basis for traditional oocyst identification. Morphologically, the sporulated oocysts were similar to those of sheep eimerian parasies; Eimeria faurei and Eimeria crandallis. PCR products from the two eimerian species detected from Sawakni and Harrie breeds were sequenced and were found to be distinct from each other with mutations at five positions. One of them clustered with E. crandallis with 99.8%-100% identity with sequences available in GenBank. E. crandallis was obtained from two Sawakni sheep and two Harrie sheep. The other sequences grouped with E. faurei with 99.8% identity with the only sequences available in GenBank. E. crandallis was detected from both Sawakni and Harrie breeds whereas E. faurei was detected only from Sawakni sheep. The findings of this study have implications for the importance of morphometric identification with advanced molecular tools to confirm the identities of sheep Eimeria species and to address the taxonomic study of this eimeriid parasite at the species level.

Highlighting the potential for crude oil bioremediation of locally isolated Cunninghamella echinulata and Mucor circinelloides.

The current investigation was carried out to assess the potential of fungi isolated from polluted soil samples in Al Jubail, Saudi Arabia, to degrade crude oil. In a minimal salt medium with 1% crude oil as the carbon source, the growth potential of various fungal isolates was examined. Among twelve fungal isolates, YS-6 and YS-10, identified as Cunninghamella echinulata and Mucor circinelloides based on multiple sequence comparisons and phylogenetic analyses, were selected as having superior crude oil degrading abilities. To the best of our knowledge, the isolated species have never been detected in polluted soil samples in the eastern province of Saudi Arabia. YS-6 and YS-10 have shown their capacity to metabolize crude oil by removing 59.7 and 78.1% of crude oil, respectively. Interestingly, they succeeded in reducing the surface tension to 41.2 and 35.9 mN/m, respectively. Moreover, the emulsification activity and hydrophobicity were determined to be 36.7, 44.9, 35.9, and 53.4%, respectively. The recovery assays included zinc sulfate, ammonium sulfate, acid precipitation, and solvent extraction techniques. All these approaches showed that the amount of biosurfactants correlates to the tested hydrocarbons. Furthermore, the enzyme activity of these two isolates generated significantly more laccase (Lac) than manganese peroxidase (MnP) and lignin peroxidase (LiP), as compared to the control. In conclusion, our study highlights new perspectives on the fungal resources found in persistently polluted terrestrial ecosystems. This knowledge will be useful for bioremediation, safe disposal of petroleum-oil contamination, and other industrial uses.

An Insight into In Vitro Antioxidant, Antimicrobial, Cytotoxic, and Apoptosis Induction Potential of Mangiferin, a Bioactive Compound Derived from Mangifera indica.

Due to their low cost, toxicity, and health risks, medicinal plants have come to be seen as useful products and sources of biologically active compounds. Mangifera indica L., a medicinal plant with a long history, has a high bioactive metabolites content. Mangiferin (C19H18O11) is primary isolated from M. indica's leaves, which has many pharmacological benefits. In this investigation, ultrasonic-assisted extraction with ethanol as the extraction solvent was applied to obtain mangiferin from a local type of M. indica leaves. HPLC was performed after a dichloromethane-ethyl acetate liquid-liquid fractionation method. Further, UV-vis, FTIR, and NMR spectroscopy were utilized to elucidate the structure. Interestingly, purified mangiferin displayed promising antimicrobial efficacy against a diverse variety of fungal and bacterial pathogens with MICs of 1.95-62.5 and 1.95-31.25 µg/mL, respectively. Time-kill patterns also showed that mangiferin had both bactericidal and fungicidal action. Furthermore, it exhibited strong radical dosage-dependent scavenging activity (IC50 = 17.6 µg/mL) compared to vitamin C (Vc, IC50 = 11.9 µg/mL), suggesting it could be developed into a viable antioxidant agent. To our delight, the IC50 values of mangiferin for the MCF-7 and HeLa cell lines were 41.2 and 44.7 µg/mL, respectively, from MTT cell viability testing, and it was less harmful when tested against the noncancerous cell line. Notably, it significantly induced cell apoptosis in MCF-7 cells by 62.2-83.4% using annexin V-FITC/PI labeling. Hence, our findings suggest that mangiferin can be used in the medical industry to create therapeutic interventions and medication delivery systems for society.

Multi-Function of a New Bioactive Secondary Metabolite Derived from Endophytic Fungus Colletotrichum acutatum of Angelica sinensis.

In the current study we assessed a new crystallized compound, 5-(1-hydroxybutyl)-4-methoxy-3-methyl-2H-pyran-2-one (C-HMMP), from the endophytic fungus Colletotrichum acutatum residing in the medicinal plant Angelica sinensis for its in vitro antimicrobial, antibiofilm, antioxidant, antimalarial, and anti-proliferative properties. The promising compound was identified as C-HMMP through antimicrobial-guided fraction. The structure of C-HMMP was unambiguously confirmed by 2D NMR and HIRS spectroscopic analysis. Antimicrobial property testing of C-HMMP showed it to be effective against a variety of pathogenic bacteria and fungi with MICs ranging from 3.9 to 31.25 µg/ml. The compound displayed excellent antibiofilm activity against C. albicans, S. aureus, and K. pneumonia. Furthermore, the antimalarial and radical scavenging activities of C-HMMP were clearly dosede-pendent, with IC50 values of 0.15 and 131.2 µg/ml. The anti-proliferative activity of C-HMMP against the HepG-2, HeLa, and MCF-7 cell lines in vitro was investigated by MTT assay, revealing notable anti-proliferative activity with IC50 values of 114.1, 90, and 133.6 µg/ml, respectively. Moreover, C-HMMP successfully targets topoisomerase I and demonstrated beneficial anti-mutagenicity in the Ames test against the reactive carcinogenic mutagen, 2-aminofluorene (2-AF). Finally, the compound inhibited the activity of α-glucosidase and α-amylase with IC50 values of 144.7 and 118.6 µg/ml, respectively. To the best of our knowledge, the identified compound C-HMMP was obtained for the first time from C. acutatum of A. sinensis, and this study demonstrated that C-HMMP has relevant biological significance and could provide better therapeutic targets against disease.

Antioxidant, Cytotoxic, and DNA Damage Protection Activities of Endophytic Fungus Pestalotiopsis neglecta Isolated from Ziziphus spina-christi Medicinal Plant.

Fungal endophytes are friendly microorganisms that colonize plants and are important in the interactions between plants and their environment. They generate valuable secondary metabolites that are valuable to both plants and humans. Endophytic fungi with bioactivities were isolated from the leaves of the medicinal plant Ziziphus spina-christi. An efficient isolate was selected and identified as Pestalotiopsis neglecta based on nucleotide sequencing of the internal transcribed spacer region (ITS 1-5.8S-ITS 2) of the 18S rRNA gene (NCBI accession number OP529850); the 564 bp had 99 to 100% similarity with P. neglecta MH860161.1, AY682935.1, KP689121.1, and MG572407.1, according to the BLASTn analysis, following preliminary phytochemical and antifungal screening. The biological activities of this fungus' crude ethyl acetate (EtOAc) extract were assessed. With an efficient radical scavenging activity against 2,2'-diphenyl-1-picrylhydrazyl and an IC50 value of 36.6 µg mL-1, P. neglecta extract has shown its potential as an antioxidant. Moreover, it displayed notable cytotoxic effects against MCF-7 (breast carcinoma, IC50 = 22.4 µg mL-1), HeLa (cervical carcinoma, IC50 = 28.9 µg mL-1) and HepG-2 (liver carcinoma, IC50 = 28.9 µg mL-1). At 10 µg mL-1, EtOAc demonstrated significant DNA protection against hydroxyl radical-induced damage. Based on FT-IR and GC-MS spectral analysis, it was detected that the EtOAc of P. neglecta product contains multiple bioactive functional groups. Subsequently, this validated the features of major different potent compounds; tolycaine, 1H-pyrazol, 1,3,5-trimethyl-, eugenol, 2,5-cyclohexadiene-1,4-dione, 2,6-bis(1,1-dimethyl), and bis(2-ethylhexyl) phthalate. Since these compounds are biologically relevant in various aspects, and distinct biological activities of fungal extract were acceptable in vitro, this suggests that endophytic fungus P. neglecta may be a viable source of bioactive natural products. This could be a good starting point for pharmaceutical applications.

Increasing atmospheric CO2 differentially supports arsenite stress mitigating impact of arbuscular mycorrhizal fungi in wheat and soybean plants.

Arbuscular mycorrhizal fungi (AMF) are beneficial for the plant growth under heavy metal stress. Such beneficial effect is improved by elevated CO2 (eCO2). However, the mechanisms by which eCO2 improves AMF symbiotic associations under arsenite (AsIII) toxicity are hardly studied. Herein, we compared these regulatory mechanisms in species from two agronomical important plant families - grasses (wheat) and legumes (soybean). AsIII decreased plant growth (i.e., 53.75 and 60.29% of wheat and soybean, respectively) and photosynthesis. It also increased photorespiration and oxidative injury in both species, but soybean was more sensitive to oxidative stress as indicated by higher H2O2 accumulation and oxidation of protein and lipid. eCO2 significantly improved AMF colonization by increasing auxin levels, which induced high carotenoid cleavage dioxygenase (CCDs) activity, particularly in soybean roots. The improved sugar metabolism in plant shoots by co-application of eCO2 and AsIII allocated more sugars to roots sequentially. Sugar accumulation in plant roots is further induced by AMF, resulting in more C skeletons to produce organic acids, which are effectively exudated into the soil to reduce AsIII uptake. Exposure to eCO2 reduced oxidative damage and this mitigation was stronger in soybean. This could be attributed to a greater reduction in photorespiration as well as a stronger antioxidant and detoxification defence systems. The grass/legume-specificity was supported by principal component analysis, which revealed that soybean was more affected by AsIII stress and more responsive to AMF and eCO2. This study provided a mechanistic understanding of the impact of AMF, eCO2 and their interaction on As-stressed grass and legume plants, allowing better practical strategies to mitigate AsIII phytotoxicity.

Anti-proliferative and apoptosis induction activities of curcumin on Leishmania major.

Leishmaniasis is a major vector-borne disease triggered by an obligate intracellular protozoan parasite of the genus Leishmania and transmitted by the bite of phlebotomine female sand flies. This parasite causes a wide range of human diseases, from localized self-healing cutaneous lesions to fatal visceral infections. The aim of this study was to investigate the cytotoxic, antiproliferative, and apoptotic effects of curcumin on Leishmania major promastigotes (MHOM/SA/84/JISH) and to assess these effects on the cell cycle of promastigotes. The MTT colorimetric assay was used to evaluate the cytotoxicity and proliferation of promastigotes. Additionally, flow cytometry was used to analyze the cell cycle. The Annexin V/propidium iodide staining technique followed by flow cytometry was used to study the cell death induced by curcumin. In this study curcumin showed a potent antileishmanial effect, exhibiting cytotoxicity against L. major promastigotes. At 80µM, the survival in curcumin treated promastigotes reached 22%; however, the median lethal concentration of curcumin (LC50) was 35µM. The drug exerted its cytotoxic effect by inducing apoptosis. Curcumin-induced cell death in promastigotes reached 82.5% at 80µM concentration. In addition, curcumin delayed the cell cycle in the S-phase inhibiting cell proliferation. Thus, curcumin was shown to be effective against L. major promastigotes. Therefore, curcumin merits further research studies to demonstrate its efficacy in treating cutaneous leishmaniasis.

Interactive Impact of Arbuscular Mycorrhizal Fungi and Elevated CO2 on Growth and Functional Food Value of Thymus vulgare.

Arbuscular mycorrhizal fungi (AMF) and elevated CO2 (eCO2) have been effectively integrated to the agricultural procedures as an ecofriendly approach to support the production and quality of plants. However, less attention has been given to the synchronous application of AMF and eCO2 and how that could affect the global plant metabolism. This study was conducted to investigate the effects of AMF and eCO2, individually or in combination, on growth, photosynthesis, metabolism and the functional food value of Thymus vulgare. Results revealed that both AMF and eCO2 treatments improved the photosynthesis and biomass production, however much more positive impact was obtained by their synchronous application. Moreover, the levels of the majority of the detected sugars, organic acids, amino acids, unsaturated fatty acids, volatile compounds, phenolic acids and flavonoids were further improved as a result of the synergistic action of AMF and eCO2, as compared to the individual treatments. Overall, this study clearly shows that co-application of AMF and eCO2 induces a synergistic biofertilization impact and enhances the functional food value of T. vulgare by affecting its global metabolism.

Global metabolic changes induced by arbuscular mycorrhizal fungi in oregano plants grown under ambient and elevated levels of atmospheric CO2.

Symbiotic plant-microorganisms interaction is a promising approach to avoid the environmental hazards of synthetic fertilizers and pesticides. Among these, arbuscular mycorrhizal fungi (AMF) are known to improve the growth and quality of many plant species; however the detailed metabolic mechanisms behind such beneficial effects are far from complete. Further, elevated levels of atmospheric CO2 (eCO2) could affect such AMF-plant association. Herein, we have investigated the individual and synchronous impact of AMF and eCO2 (620 ppm) on nutrient uptake, growth, photosynthesis, respiration, and levels of primary and secondary metabolites in oregano (Oreganum vulgare), an economically important herbal plant. Enhanced AMF colonization rate and a better mycelial growth were observed in roots of oregano grown under eCO2. Both AMF and eCO2 treatments significantly enhanced the growth and photosynthesis of oregano plants, however much improvements were observed by their synchronous application. eCO2 further increased the AMF-induced dark respiration and accumulation of macro and microelements. Hierarchical clustering analysis of individual primary and secondary metabolites revealed a metabolite-dependent response toward AMF and eCO2. The synchronous application of AMF and eCO2 resulted in promoted accumulation of the majority of the detected sugars, organic acids, amino acids, unsaturated fatty acids, phenolic acids and flavonoids, as compared with the sole treatments. Moreover, AMF and eCO2 acted synergistically in improving the antioxidant capacity and anti-lipid peroxidation activity of oregano. Therefore, this study suggests that AMF treatment induces a global metabolic change in oregano, the effect that is strengthened under eCO2.

Bacterial contamination and health risks of drinking water from the municipal non-government managed water treatment plants.

Water quality and bacterial contamination from 18 drinking water municipal plants in three locations at Giza governorate were investigated. The average total count of bacteria detected after four stages of treatments in the investigated plants was 32 CFU/1 mL compared to 2330 cfu/mL for raw water, with a reduction percentage of 98.6. Although there is a relatively high removal percent of bacterial contamination from the water sources, however, several bacterial pathogens were identified in the produced water prepared for drinking including Enterococcus faecalis, Escherichia coli, Pseudomonas aeruginosa, and Shigella spp. After 3 days of water incubation at 30 °C, the amount of bacterial endotoxins ranged from 77 to 137 ng/mL in the water produced from the municipal plants compared to 621-1260 ng/mL for untreated water. The main diseases reported from patients attending different clinics and hospitals during summer 2014 at the surveyed locations and assuredly due to drinking water from these plants indicated that diarrheas and gastroenteritis due to E. coli and Campylobacter jejuni constituted 65.7% of the total patients followed by bacillary dysentery or shigellosis due to Shigella spp. (7.9%) and cholera due to Vibrio cholera (7.2%). There was an increase in serum aspartate aminotransferase (AST), alanine aminotransferase (ALT), and alkaline phosphatase (ALP) as well as urea and creatinine values of guinea pigs consuming water produced from the non-governmental plants for 6 months indicating remarkable liver and kidney damages. Histological sections of liver and kidney from the tested animal revealed liver having ballooning degeneration of hepatocytes and distortion and fragmentation of the nuclei, while the section of the kidney showed irregularly distributed wrinkled cells, degenerated Bowman's capsule, congested blood vessels, and inflammatory cells.

Biosynthesis and characterization of silver nanoparticles produced by Pleurotus ostreatus and their anticandidal and anticancer activities.

The biosynthesis of nanoparticles has received increasing interest because of the growing need to develop safe, cost-effective and environmentally friendly technologies for the synthesis of nano-materials. In this study, silver nanoparticles (AgNPs) were synthesized using a reduction of aqueous Ag(+) ions with culture supernatant from Pleurotus ostreatus. The bioreduction of AgNPs was monitored by ultra violet-visible spectroscopy and the obtained AgNPs were characterized by transmission electron microscopy (TEM) and Fourier transform infrared spectroscopy techniques. TEM studies showed the size of the AgNPs to be in the range of 4-15 nm. The formation of AgNPs might be an enzyme-mediated extracellular reaction process. Furthermore, the antifungal effect of AgNPs against Candida albicans as compared with commercially antifungal drugs was examined. The effect of AgNPs on dimorphic transition of C. albicans was tested. The anticancer properties of AgNPs against cells (MCF-7) were also evaluated. AgNPs caused a significant decrease in cell viability of an MCF-7 cell line (breast carcinoma). Exposure of MCF-7 cells with AgNPs resulted in a dose-dependent increase in cell growth inhibition varying from 5 to 78 % at concentrations in the range of 10-640 µg ml(-1). The present study demonstrated that AgNPs have potent antifungal, antidimorphic, and anticancer activities. The current research opens a new avenue for the green synthesis of nano-materials.