Dissemination of mcr-1 and ß-lactamase genes among Pseudomonas aeruginosa: molecular characterization of MDR strains in broiler chicks and dead-in-shell chicks infections.

OBJECTIVES: Pseudomonas aeruginosa (P. aeruginosa) is one of the most serious pathogens implicated in antimicrobial resistance, and it has been identified as an ESKAPE along with other extremely significant multidrug resistance pathogens. The present study was carried out to explore prevalence, antibiotic susceptibility phenotypes, virulence-associated genes, integron (int1), colistin (mcr-1), and ß-lactamase resistance' genes (ESBls), as well as biofilm profiling of P. aeruginosa isolated from broiler chicks and dead in-shell chicks. DESIGN: A total of 300 samples from broiler chicks (n = 200) and dead in-shell chicks (n = 100) collected from different farms and hatcheries located at Mansoura, Dakahlia Governorate, Egypt were included in this study. Bacteriological examination was performed by cultivation of the samples on the surface of both Cetrimide and MacConkey's agar. Presumptive colonies were then subjected to biochemical tests and Polymerase Chain Reaction (PCR) targeting 16S rRNA. The recovered isolates were tested for the presence of three selected virulence-associated genes (lasB, toxA, and exoS). Furthermore, the retrieved isolates were subjected to phenotypic antimicrobial susceptibility testing by Kirby-Bauer disc diffusion method as well as phenotypic detection of ESBLs by both Double Disc Synergy Test (DDST) and the Phenotypic Confirmatory Disc Diffusion Test (PCDDT). P. aeruginosa isolates were then tested for the presence of antibiotic resistance genes (ARGs): int1, mcr-1, and ESBL genes (OXA-10, OXA-2, VEB-1, SHV, TEM, and CTX-M). Additionally, biofilm production was examined by the Tube Adherent method (TA) and Microtiter Plate assay (MTP). RESULTS: Fifty -five isolates were confirmed to be P. aeruginosa, including 35 isolates from broiler chicks and 20 isolates from dead in-shell chicks. The three tested virulence genes (lasB, toxA, and exoS) were detected in all isolates. Antibiogram results showed complete resistance against penicillin, amoxicillin, ceftriaxone, ceftazidime, streptomycin, erythromycin, spectinomycin, and doxycycline, while a higher sensitivity was observed against meropenem, imipenem, colistin sulfate, ciprofloxacin, and gentamicin. ESBL production was confirmed in 12 (21.8%) and 15 (27.3%) isolates by DDST and PCDDT, respectively. Antibiotic resistance genes (ARGs): int1, mcr-1, and ESBL genes (OXA-10, SHV, TEM, and CTX-M), were detected in 87.3%, 18.2%, 16.4%, 69.1%, 72.7%, and 54.5% of the examined isolates respectively, whereas no isolate harbored the OXA-2 or VEB-1 genes. Based on the results of both methods used for detection of biofilm formation, Kappa statistics [kappa 0.324] revealed a poor agreement between both methods. CONCLUSIONS: the emergence of mcr-1 and its coexistence with other resistance genes such as ß-lactamase genes, particularly blaOXA-10, for the first time in P. aeruginosa from young broiler chicks and dead in-shell chicks in Egypt pose a risk not only to the poultry industry but also to public health.

Response Surface Methodology (RSM) approach to formulate and optimize the bilayer combination tablet of Tamsulosin and Finasteride.

An orally administered bilayer tablet with Tamsulosin (TAM) as the sustained release (SR) and Finasteride (FIN) as immediate release (IR) was manufactured. A response surface methodology was employed to formulate bilayer tablets with individual release layers, i.e., sustained and immediate release (SR and IR). Independent variables selected in both cases comprise hydroxypropyl methylcellulose (HPMC) as SR polymer, and avicel PH102 in the inner layer while Triacetin and talc in the outer layer, respectively. Tablets were prepared by direct compression, a total of 11 formulations were prepared for inner layer TAM, and 9 formulations for outer layer FIN were designed; these formulations were evaluated for hardness, friability, thickness, %drug content, and %drug release. A central composite design was employed in response surface methodology to design and optimize the formulation. The percentage of drug released was evaluated by in-vitro USP dissolution method of optimized formulation for 0.5, 2, and 6 hrs, and results were 24.63, 52.96, and 97.68 %, respectively. Drug release data was plotted in various kinetic models using a D.D solver, where drug release was first order that is concentration dependent and was best explained by Korsmeyer-Peppa kinetics, as the highest linearity was observed (R2 = 0.9693). However, a very close relationship was also noted with Higuchi kinetics (R2 = 0.9358). The mechanism of drug release was determined through the Korsmeyer model, and exponent "n" was found to be 0.4, indicative of an anomalous diffusion mechanism or diffusion coupled with erosion.

Biochemical and phenological characterization of diverse wheats and their association with drought tolerance genes.

Drought is one of the most important wheat production limiting factor, and can lead to severe yield losses. This study was designed to examine the effect of drought stress on wheat physiology and morphology under three different field capacities (FC) viz. 80% (control), 50% (moderate) and 30% (severe drought stress) in a diverse collection of wheat germplasm including cultivars, landraces, synthetic hexaploid and their derivatives. Traits like grain weight, thousand grain weight and biomass were reduced by 38.23%, 18.91% and 26.47% respectively at 30% FC, whereas the reduction rate for these traits at 50% FC were 19.57%, 8.88% and 18.68%. In principal component analysis (PCA), the first two components PC1 and PC2 accounted for 58.63% of the total variation and separated the cultivars and landraces from synthetic-based germplasm. Landraces showed wide range of phenotypic variations at 30% FC compared to synthetic-based germplasm and improved cultivars. However, least reduction in grain weight was observed in improved cultivars which indicated the progress in developing drought resilient cultivars. Allelic variations of the drought-related genes including TaSnRK2.9-5A, TaLTPs-11, TaLTPs-12, TaSAP-7B-, TaPPH-13, Dreb-B1 and 1fehw3 were significantly associated with the phenological traits under drought stress in all 91 wheats including 40 landraces, 9 varieties, 34 synthetic hexaploids and 8 synthetic derivatives. The favorable haplotypes of 1fehw3, Dreb-B1, TaLTPs-11 and TaLTPs-12 increased grain weight, and biomass. Our results iterated the fact that landraces could be promising source to deploy drought adaptability in wheat breeding. The study further identified drought tolerant wheat genetic resources across various backgrounds and identified favourable haplotypes of water-saving genes which should be considered to develop drought tolerant varieties.

A Novel Based Synthesis of Silver/Silver Chloride Nanoparticles from Stachys emodi Efficiently Controls Erwinia carotovora, the Causal Agent of Blackleg and Soft Rot of Potato.

In recent years, the biological synthesis of silver nanoparticles has captured researchers' attention due to their unique chemical, physical and biological properties. In this study, we report an efficient, nonhazardous, and eco-friendly method for the production of antibacterial silver/silver chloride nanoparticles utilizing the leaf extract of Stachys emodi. The synthesis of se-Ag/AgClNPs was confirmed using UV-visible spectroscopy, DPPH free radical scavenging activity, Fourier transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM), and X-ray diffraction (XRD). An intense peak absorbance was observed at 437 nm from the UV-visible analysis. The Stachys emodi extract showed the highest DPPH scavenging activity (89.4%). FTIR analysis detected various bands that indicated the presence of important functional groups. The SEM morphological study revealed spherical-shaped nanoparticles having a size ranging from 20 to 70 nm. The XRD pattern showed the formation of a spherical crystal of NPs. The antibacterial activity performed against Erwinia carotovora showed the maximum inhibition by centrifuged silver nanoparticles alone (se-Ag/AgClNPs) and in combination with leaf extract (se-Ag/AgClNPs + LE) and leaf extract (LE) of 98%, 93%, and 62% respectively. These findings suggested that biosynthesized NPs can be used to control plant pathogens effectively.

Design, synthesis and cytotoxic activity of molecular hybrids based on quinolin-8-yloxy and cinnamide hybrids and their apoptosis inducing property.

The present work aims at design and synthesis of a congeneric series of small hybrids 5 and 6a-i featuring the privileged quinoline scaffold tethered with 2-(arylamido)cinnamide moiety as potential anticancer tubulin polymerization inhibitors. Most of the synthesized hybrids 5 and 6a-i significantly inhibited the growth of the HepG2 cell line, with IC50 ranged from 2.46 to 41.31 µM. In particular, 2-(3,4,5-trimethoxybenzamido)-4-methoxycinnamide-quinoline hybrid 6e displayed potent IC50 value toward the examined cell line, and hence chosen for further mechanistic investigations. It is noteworthy that the antiproliferative action of compound 6e highly correlated well with its ability to inhibit tubulin polymerization. In addition, the most potent hybrid 6e demonstrated a significant modification in the cellular cycle distribution, in addition to provoke of apoptotic death within the tested HepG2 cell line. Furthermore, the mechanistic approach was confirmed by a substantial upregulation in the quantity of active caspase 9 by 5.81-fold relative to untreated control cells.

Synthesis and biological research of new imidazolone-sulphonamide-pyrimidine hybrids as potential EGFR-TK inhibitors and apoptosis-inducing agents.

Development of new effective EGFR-targeted antitumor agents is needed because of their clinical significance. A new series of imidazolone-sulphonamide-pyrimidine hybrids was designed and synthesized as modified analogs of some reported EGFR inhibitors. The cytotoxic activity of all the synthesized hybrids was investigated against the breast MCF-7 cancerous cell line using doxorubicin (Dox) as a positive control. 4-(Furan-2-ylmethylene)imidazolone-sulphonamide-pyrimidine 6b had the best potent activity against MCF-7 cells with IC50 result of 1.05 µM, which was better than Dox (IC50 = 1.91 µM). In addition, mechanistic studies revealed the ability of compounds 5g, 5h and 6b to inhibit EGFR kinase. Cell cycle analysis revealed that compound 6b can halt MCF-7 cells at the G1 phase with a concomitant decrease in cellular percentage at the S and G2/M phases. This compound produced a noticeable rise in the proportion of apoptotic cells with regard to the untreated control. Furthermore, the effects of hybrid 6b on the expression levels of pro-apoptotic Bax and pro-survival Bcl2 were assessed. The results showed that this compound upregulated the level of Bax expression as well as declined the expression value of Bcl-2 with regard to the untreated control.

Exploring Bacillus mycoides PM35 efficacy in enhancing rice (Oryza sativa L.) response to different types of microplastics through gene regulation and cellular fractionation.

Soil contamination with microplastics (MPs) is a persistent threat to crop production worldwide. With a wide range of MP types, including polystyrene (PS), polyvinyl chloride (PVC) and polyethylene (PE), contaminating our environment, it is important to understand their impact on agricultural productivity. The present study was conducted to investigate the effects of different types of MPs (PS, PVC and PE) on various aspects of plant growth. Specifically, we examined growth and biomass, photosynthetic pigments, gas exchange attributes, oxidative stress responses, antioxidant compound activity (both enzymatic and non-enzymatic), gene expression, proline metabolism, the AsA-GSH cycle and cellular fractionation and nutritional status, in different parts of rice (Oryza sativa L.) seedlings, which were also exposed to plant growth promoting rhizobacteria (PGPR), i.e. Bacillus mycoides PM35, i.e. 20 µL. The research outcomes indicated that the different types of MPs in the soil notably reduced plant growth and biomass, photosynthetic pigments and gas exchange attributes. However, MP stress also induced oxidative stress in the roots and shoots of the plants by increasing malondialdehyde (MDA), hydrogen peroxide (H2O2) and electrolyte leakage (EL) which also induced increased compounds of various enzymatic and non-enzymatic antioxidants and also the gene expression. Furthermore, a significant increase in proline metabolism, the AsA-GSH cycle, and the fractionations of cellular components was observed. Although the application of B. mycoides PM35 showed a significant increase in plant growth and biomass, gas exchange characteristics, enzymatic and non-enzymatic compounds and their gene expression and also decreased oxidative stress. In addition, the application of B. mycoides PM35 enhanced cellular fractionation and decreased the proline metabolism and AsA-GSH cycle in O. sativa plants. These results open new insights for sustainable agriculture practices and hold immense promise in addressing the pressing challenges of MP contamination in agricultural soils.

In vitro anti-breast cancer study of hybrid cinnamic acid derivatives bearing 2-thiohydantoin moiety.

Aim: To synthesize new hybrid cinnamic acids (10a, 10b and 11) and ester derivatives (7, 8 and 9) and investigate their anti-breast cancer activities. Materials & methods: Compounds 7-11 were evaluated (in vitro) for their cytotoxic activities against the MCF-7 cell line. A flow cytometry examination was performed. Protein levels of nuclear factor erythroid 2-related factor 2 (Nrf2), topoisomerase II and caspase-9 were measured by qRT-PCR. Molecular docking studies were conducted. Results: Several components were discovered to be active, mainly component 11, which induced arrest in the cell cycle at phase S, greatly decreased the expression of Nrf2 and topoisomerase II; and upregulated the expression of caspase-9. Conclusion: The newly thiohydantoin-cinnamic acid hybrids can contribute to creating promising candidates for cancer drugs.

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Evaluation of novel synthesized thiazole derivatives as potential aromatase inhibitors against breast cancer.

Background: 4-Methylacetophenone is used in the preparation of starting materials, 4-methylphenacyle bromide (2) and 4-methylacetophenone thiosemicarbazole (3). Results: Several novel 2,4-disubstituted-1,3-thiazole analogues were obtained via the treatment of starting materials with 4-methylphenacyl bromide, acetyl chloride, aromatic aldehydes and bromination providing thiazole derivatives 5-8 respectively. Conclusion: Compounds 5-8 were investigated for their cytotoxic activity on MCF-7 and normal breast cells. Active compounds were found and in contrast to staurosporine, compound 8 displayed the most potent cytotoxic action that showed a strong inhibitory effect (aromatase) and (protein tyrosine kinase) enzymes, proving that the novel thiazole derivatives promoted the effective anticancer drug candidates.

Hyperglycemia-induced oxidative stress and epigenetic regulation of ET-1 gene in endothelial cells.

Introduction: Hyperglycemia-induced endothelial dysfunction and the subsequent increase of oxidative stress could lead to aberrant regulation of various genes which are responsible for a range of functions. This study aims to find out how hyperglycemia affect oxidative stress and then the expression and methylation of endothelin 1 (ET-1) gene in in human umbilical vein endothelial cells (HUVEC). Methods: Cells were cultured in growth medium and exposed to low and high glucose concentrations to mimic normal and diabetic condition respectively. Computational analysis were performed using UCSC genome browser and eukaryotic promoter database (EPD). The expression of ET-1 gene was investigated by real time PCR. Cytotoxicity and oxidative stress were determined by MTT and DCFH-DA assays respectively. Promoter methylation was assessed by the bisulfite sequencing method. Results: DCFH-DA assay showed that hyperglycemia can significantly increase the regulation of reactive oxygen species synthesis. The relative expression of ET-1 gene was increased due to exposure to high glucose concentration. MTT assay revealed reduced viability of cells due to the glucose induced damage. Methylation analysis revealed hypomethylation of the promoter of ET-1 however the difference was not significant. Out of 175 CpGs at 25 CpG sites, only 36 CpGs were methylated (20.5% methylation) in cell treated with normal glucose. Upon exposure to high glucose only 30 CpGs were methylated in 175 CpGs at 25 CpG sites (17.1% methylation). Discussion: Our study concludes a significantly high expression of ET-1 gene in response to high glucose exposure in HUVECs. It also reports that hyperglycemic condition leads to elevated oxidative stress. No significant change was found in methylation when cells were treated with high and low glucose concentrations.

Shelf-life extension of Fragaria × ananassa Duch. using selenium nanoparticles synthesized from Cassia fistula Linn. leaves.

Fragaria × ananassa Duch. (Strawberry) fruit is susceptible to postharvest diseases, thus decrease in quality attributes, such as physiological and biochemical properties leads to decrease in shelf life. The objective of the present study was to check the effect of Selenium NP's and packaging conditions on the shelf life of strawberry (Fragaria × ananassa Duch) fruits. The shelf life was observed with 4 days intervals and examined for characteristics such as physiological weight loss, moisture content, percentage decay loss, peroxidase, catalase, and DPPH radical scavenging. The quality change of postharvest Fragaria × ananassa Duch. was monitored by the application of selenium nanoparticles (T1 plant extract in 10 mM salt solution, T2 plant extract in 30 mM salt solution, T3 plant extract in 40 mM salt solution, T4 distilled water; control) in different packaging materials (plastic bags, cardboard, and brown paper) at different storage conditions (6°C and 25°C). 10 mM, 20 mM, and 30 mM solution of sodium selenite salt, prepared from 1 M stock solution. Selenium nanoparticles were synthesized using Cassia fistula L. extract and sodium selenite salt solution. Polyvinyl alcohol (PVA) was used as a stabilizer. The nanoparticles were characterized through UV-visible spectroscopy and X-Ray diffractometer (XRD). It was observed that the strawberry Fragaria × ananassa Duch. Treated with T1 (CFE and 10 mM salt solution) stored in plastic packaging at ±6°C showed the best physiological parameters and hence the treatment is recommended for storage without affecting the quality of strawberry fruit up to 16 days.

Lectin-Fortified Cationic Copper Sulfide Nanoparticles Gain Dual Targeting Capabilities to Treat Carbapenem-Resistant Acinetobacter baumannii Infection.

Targeted drug delivery maximizes the chance to combat infection caused by drug-resistant pathogens. Herein, lectin-fortified cationic copper sulfide (cCuS) nanoparticles were suggested for targeted adhesion to bacterial membranes and to enforce bacterial death. Jacalin, a lectin from jackfruit seed, was conjugated to fluorescein isothiocyanate (FITC), and its ability to recognize bacterial cell surface glycans was demonstrated. Jacalin formed a noncovalent complex with cCuS, which was investigated by fluorescence quenching measurements. The data revealed that jacalin-cCuS (JcCuS) had a good affinity with an association constant K a of 2.27 (± 0.28) × 104 M-1. The resultant JcCuS complex displayed excellent anti-infective activity against carbapenem-resistant Acinetobacter baumannii (CRAB). The minimum inhibitory concentration (MIC) of cCuS was 62.5 µM, which was 2-fold lower than that of the broad-spectrum antibiotic ciprofloxacin. Interestingly, the MIC of JcCuS was reduced to 15.63 µM, which was attributed to jacalin fortification. The mechanistic study unveiled that JcCuS affected the membrane integrity, depolarized the inner membrane, and produced excess reactive oxygen species to combat CRAB at a lower concentration compared to cCuS. A. baumannii formed a biofilm more readily, which played a critical role in pathogenesis and resistance in clinical settings. JcCuS (3.91 µM) displayed stronger antibiofilm activity without affecting the metabolic viability of CRAB. Microscopy analyses confirmed the inhibition of biofilm formation and disruption of the mature biofilm upon treatment with JcCuS. Furthermore, JcCuS hindered pellicle formation and inhibited the biofilm-associated virulence factor of CRAB such as exopolysaccharide, cell surface hydrophobicity, swarming, and twitching mobility. The anti-infective potential of JcCuS was demonstrated by rescuing CRAB-infected zebrafish. The reduction in pathogen proliferation in muscle tissues was observed in the treated group, and the fish recovered from the infection and was restored to normal life within 12 h. The findings illustrate that lectin fortification offers a unique advantage in enhancing the therapeutic potential of antimicrobials against human pathogens of critical priority worldwide.

Wild Vicia Species Possess a Drought Tolerance System for Faba Bean Improvement.

Faba bean (Vicia faba L.), a drought-sensitive crop, is drastically affected by drought stresses compromising its growth and yield. However, wild relatives of faba bean are considered a reservoir of potential genetic resources for tolerance against abiotic stresses. This study was conducted to characterize wild relatives of faba bean for identification of a specific tolerance system required for its improvement against drought stress. The study focused on physiological, biochemical, and anatomical responses of wild Vicia species under drought stress conditions. The experiment was carried out under various levels of drought stress imposed through different field capacities (FC) which included 80% FC ie (well-watered condition), 55% FC (moderate stress), and 30% FC (severe stress). When compared to plants grown in a control environment, drought stress significantly reduced the studied physiological attributes including soluble sugars (21.3% and 15.8%), protein contents (14.7 and 14.6%), and chlorophyll (8.4 and 28.6%) under moderate (55% FC) and severe drought stress (30% FC), respectively. However, proline content increased by 20.5% and 27.6%, peroxidase activity by 48.5% and 57.1%, and superoxide dismutase activity by 72.6% and 64.8% under moderate and severe stress, respectively. The studied anatomical attributes were also affected under drought stress treatments, including diameter of stem xylem vessels (9.1% and 13.7%), leaf lower epidermal thickness (8.05% and 13.34%), and leaf phloem width (5.3% and 10.1%) under moderate and severe stress, respectively. Wild Vicia spp. showed better tolerance to water-deficit conditions as compared to cultivated Vicia L. The observed potential diversity for drought tolerance in wild Vicia spp. may assist in improvement of faba bean and may also help in understanding the mechanisms of adaptations in drought-prone environments.

Molecular and Physiological Evaluation of Bread Wheat (Triticum aestivum L.) Genotypes for Stay Green under Drought Stress.

Water availability is considered as the main limiting factor of wheat growth illuminating the need of cultivars best adapted to drought situations for better wheat production and yield. Among these, the stay-green trait is thought to be related to the ability of wheat plants to maintain photosynthesis and CO2 assimilation, and a detailed molecular understanding of this trait may help in the selection of high-yielding, drought-tolerant wheats. The current study, therefore, evaluated the physiological responses of the selected wheat genotypes under pot-induced water stress conditions through different field capacities. The study also focused on exploring the molecular mechanisms involved in drought tolerance conferred due to the stay-green trait by studying the expression pattern of the selected PSI-associated light-harvesting complex I (LHC1) and PSII-associated LHCII gene families related to pigment-binding proteins. The results revealed that the studied traits, including relative water content, membrane stability index and chlorophyll, were variably and negatively affected, while the proline content was positively enhanced in the studied wheats under water stress treatments. Molecular diagnosis of the selected wheat genotypes using the expression profile of 06 genes, viz. TaLhca1, TaLhca2, TaLhca3, TaLhcb1, TaLhcb4 and TaLhcb6 that encodes for the LHCI and LHCII proteins, indicated variable responses to different levels of drought stress. The results obtained showed the relation between the genotypes and the severity of the drought stress condition. Among the studied genotypes, Chirya-1 and SD-28 performed well with a higher level of gene expression under drought stress conditions and may be used in genetic crosses to enrich the genetic background of common wheat against drought stress.