Encapsulation of E. coli phage ZCEC5 in chitosan-alginate beads as a delivery system in phage therapy

© 2019, The Author(s). Bacteriophages can be used successfully to treat pathogenic bacteria in the food chain including zoonotic pathogens that colonize the intestines of farm animals. However, harsh gastric conditions of low pH and digestive enzyme activities affect phage viability, and accordingly reduce their effectiveness. We report the development of a natural protective barrier suitable for oral administration to farm animals that confers acid stability before functional release of bead-encapsulated phages. Escherichia coli bacteriophage ZSEC5 is rendered inactive at pH 2.0 but encapsulation in chitosan–alginate bead with a honey and gelatin matrix limited titer reductions to 1log10PFUmL−1. The encapsulated phage titers were stable upon storage in water but achieved near complete release over 4–5h in a simulated intestinal solution (0.1% bile salt, 0.4% pancreatin, 50mM KH2PO4 pH 7.5) at 37°C. Exposure of E. coli O157:H7 to the bead-encapsulated phage preparations produced a delayed response, reaching a maximal reductions of 4.2 to 4.8log10CFUmL−1 after 10h at 37°C under simulated intestinal conditions compared to a maximal reduction of 5.1log10CFUmL−1 at 3h for free phage applied at MOI = 1. Bead-encapsulation is a promising reliable and cost-effective method for the functional delivery of bacteriophage targeting intestinal bacteria of farm animals

The green production of eco-friendly silver with cobalt ferrite nanocomposite using Citrus limon extract

This paper reveals a novel and greener strategy to biosynthesize Cobalt-Ferrite-Silver trimetallic nanoparticles (Co-Fe-AgNPs) using the Citrus limon extract as a bio-reducing agent. Depending on the Transmission and Scanning Electron Microscopy (TEM and SEM), the prepared nanocomposites had a relatively small size with an average of 20 nm with a spherical shape. Moreover, The Energy Dispersive X-ray (EDX) showed the distribution of elements within the nanoparticles with various concentrations of C, K, S, O, Fe, Co and Ag. Finally, the potential ability of nanoparticles to combat the bacteria and biofilm formation was studied in-vitro. For example, a 125 µM nanocomposite is needed to kill Salmonella, and 62.5 µM inhibits its growth. Moreover, 30 µM is enough to inhibit biofilm formation. Furthermore, the anticancer activity was evaluated against MCF7 and HEPG2 cell lines with IC50 of 35.5 and 43.5 μg/mL, respectively. The findings suggest that Co-Fe-AgNPs could be a potential antibacterial and anticancer agent

The Synergistic Effect of Biosynthesized Silver Nanoparticles and Phage ZCSE2 as a Novel Approach to Combat Multidrug-Resistant Salmonella enterica

The emergence and evolution of antibiotic-resistant bacteria is considered a public health concern. Salmonella is one of the most common pathogens that cause high mortality and morbidity rates in humans, animals, and poultry annually. In this work, we developed a combination of silver nanoparticles (AgNPs) with bacteriophage (phage) as an antimicrobial agent to control microbial growth. The synthesized AgNPs with propolis were characterized by testing their color change from transparent to deep brown by transmission electron microscopy (TEM) and Fourier-Transform Infrared Spectroscopy (FTIR). The phage ZCSE2 was found to be stable when combined with AgNPs. Both minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were evaluated for AgNPs, phage, and their combination. The results indicated that MIC and MBC values were equal to 23 µg/mL against Salmonella bacteria at a concentration of 107 CFU/mL. The combination of 0.4× MIC from AgNPs and phage with Multiplicity of Infection (MOI) 0.1 showed an inhibitory effect. This combination of AgNPs and phage offers a prospect of nanoparticles with significantly enhanced antibacterial properties and therapeutic performance

Isolation and Characterization of Bacteriophage ZCSE6 against Salmonella spp.: Phage Application in Milk

Food safety is very important in the food industry as most pathogenic bacteria can cause food-borne diseases and negatively affect public health. In the milk industry, contamination with Salmonella has always been a challenge, but the risks have dramatically increased as almost all bacteria now show resistance to a wide range of commercial antibiotics. This study aimed to isolate a bacteriophage to be used as a bactericidal agent against Salmonella in milk and dairy products. Here, phage ZCSE6 has been isolated from raw milk sample sand molecularly and chemically characterized. At different multiplicities of infection (MOIs) of 0.1, 0.01, and 0.001, the phage–Salmonella interaction was studied for 6 h at 37 °C and 24 h at 8 °C. In addition, ZCSE6 was tested against Salmonella contamination in milk to examine its lytic activity for 3 h at 37 °C. The results showed that ZCSE6 has a small genome size (<48.5 kbp) and belongs to the Siphovirus family. Phage ZCSE6 revealed a high thermal and pH stability at various conditions that mimic milk manufacturing and supply chain conditions. It also demonstrated a significant reduction in Salmonella concentration in media at various MOIs, with higher bacterial eradication at higher MOI. Moreover, it significantly reduced Salmonella growth (MOI 1) in milk, manifesting a 1000-fold decrease in bacteria concentration following 3 h incubation at 37 °C. The results highlighted the strong ability of ZCSE6 to kill Salmonella and control its growth in milk. Thus, ZCSE6 is recommended as a biocontrol agent in milk to limit bacterial growth and increase the milk shelf-life

Morphological, biological, and genomic characterization of Klebsiella pneumoniae phage vB_Kpn_ZC2

Abstract Background Bacteriophages (phages) are one of the most promising alternatives to traditional antibiotic therapies, especially against multidrug-resistant bacteria. Klebsiella pneumoniae is considered to be an opportunistic pathogen that can cause life-threatening infections. Thus, this study aims at the characterization of a novel isolated phage vB_Kpn_ZC2 (ZCKP2, for short). Methods The phage ZCKP2 was isolated from sewage water by using the clinical isolate KP/08 as a host strain. The isolated bacteriophage was purified and amplified, followed by testing of its molecular weight using Pulse-Field Gel Electrophoresis (PFGE), transmission electron microscopy, antibacterial activity against a panel of other Klebsiella pneumoniae hosts, stability studies, and whole genome sequencing. Results Phage ZCKP2 belongs morphologically to siphoviruses as indicated from the Transmission Electron Microscopy microgram. The Pulsed Field Gel Electrophoresis and the phage sequencing estimated the phage genome size of 48.2 kbp. Moreover, the absence of lysogeny-related genes, antibiotic resistance genes, and virulence genes in the annotated genome suggests that phage ZCKP2 is safe for therapeutic use. Genome-based taxonomic analysis indicates that phage ZCKP2 represents a new family that has not been formally rated yet. In addition, phage ZCKP2 preserved high stability at different temperatures and pH values (-20 − 70 °C and pH 4 – 9). For the antibacterial activity, phage ZCKP2 maintained consistent clear zones on KP/08 bacteria along with other hosts, in addition to effective bacterial killing over time at different MOIs (0.1, 1, and 10). Also, the genome annotation predicted antibacterial lytic enzymes. Furthermore, the topology of class II holins was predicted in some putative proteins with dual transmembrane domains that contribute significantly to antibacterial activity. Phage ZCKP2 characterization demonstrates safety and efficiency against multidrug-resistant K. pneumoniae, hence ZCKP2 is a good candidate for further in vivo and phage therapy clinical applications

The Lytic Activity of Bacteriophage ZCSE9 against <i>Salmonella enterica</i> and Its Synergistic Effects with Kanamycin

Salmonella, the causative agent of several diseases in humans and animals, including salmonellosis, septicemia, typhoid fever, and fowl typhoid, poses a serious threat to global public health and food safety. Globally, reports of therapeutic failures are increasing because of the increase in bacterial antibiotic resistance. Thus, this work highlights the combined phage–antibiotic therapy as a promising approach to combating bacterial resistance. In this manner, the phage ZCSE9 was isolated, and the morphology, host infectivity, killing curve, combination with kanamycin, and genome analysis of this phage were all examined. Morphologically, phage ZCSE9 is a siphovirus with a relatively broad host range. In addition, the phage can tolerate high temperatures until 80 °C with one log reduction and a basic environment (pH 11) without a significant decline. Furthermore, the phage prevents bacterial growth in the planktonic state, according to the results of the time-killing curve. Moreover, using the phage at MOI 0.1 with kanamycin against five different Salmonella serotypes reduces the required antibiotics to inhibit the growth of the bacteria. Comparative genomics and phylogenetic analysis suggested that phage ZCSE9, along with its close relatives Salmonella phages vB_SenS_AG11 and wksl3, belongs to the genus Jerseyvirus. In conclusion, phage ZCSE9 and kanamycin form a robust heterologous antibacterial combination that enhances the effectiveness of a phage-only approach for combating Salmonella

Exploring Salinity Tolerance Mechanisms in Diverse Wheat Genotypes Using Physiological, Anatomical, Agronomic and Gene Expression Analyses

Salinity is a widespread abiotic stress that devastatingly impacts wheat growth and restricts its productivity worldwide. The present study is aimed at elucidating biochemical, physiological, anatomical, gene expression analysis, and agronomic responses of three diverse wheat genotypes to different salinity levels. A salinity treatment of 5000 and 7000 ppm gradually reduced photosynthetic pigments, anatomical root and leaf measurements and agronomic traits of all evaluated wheat genotypes (Ismailia line, Misr 1, and Misr 3). In addition, increasing salinity levels substantially decreased all anatomical root and leaf measurements except sclerenchyma tissue upper and lower vascular bundle thickness compared with unstressed plants. However, proline content in stressed plants was stimulated by increasing salinity levels in all evaluated wheat genotypes. Moreover, Na+ ions content and antioxidant enzyme activities in stressed leaves increased the high level of salinity in all genotypes. The evaluated wheat genotypes demonstrated substantial variations in all studied characters. The Ismailia line exhibited the uppermost performance in photosynthetic pigments under both salinity levels. Additionally, the Ismailia line was superior in the activity of superoxide dismutase (SOD), catalase activity (CAT), peroxidase (POX), and polyphenol oxidase (PPO) enzymes followed by Misr 1. Moreover, the Ismailia line recorded the maximum anatomical root and leaf measurements under salinity stress, which enhanced its tolerance to salinity stress. The Ismailia line and Misr 3 presented high up-regulation of H+ATPase, NHX2 HAK, and HKT genes in the root and leaf under both salinity levels. The positive physiological, anatomical, and molecular responses of the Ismailia line under salinity stress were reflected on agronomic performance and exhibited superior values of all evaluated agronomic traits

The Potential Utility of Circulating Oncofetal H19 Derived miR-675 Expression versus Tissue lncRNA-H19 Expression in Diagnosis and Prognosis of HCC in Egyptian Patients

Hepatocellular carcinoma (HCC) is one of the most common cancers worldwide. Interestingly, lncRNA-H19 acts independently in HCC and influences miR-675 expressions. We aimed to assess the potential utility of tissue lncRNA-H19 versus miR-675 expressions as a non-invasive biomarker for HCC diagnosis and prognosis in Egyptian patients. Ninety-one HCC patients and 91 controls included in this study were investigated for expression of lncRNA-H19 and miR675 using RT-qPCR. Our results showed that the expression of lncRNA-H19 and microRNA-675 were higher in patients than in controls (p p p < 0.001, 0.001, respectively). Oncofetal H19-derived miR-675 expression could be considered a potential noninvasive diagnostic and prognostic biomarker, outstanding the performance of the expression of tissue lncRNA-H19 for HCC