Inhibition Mechanism of Methicillin-Resistant Staphylococcus aureus by Zinc Oxide Nanorods via Suppresses Penicillin-Binding Protein 2a.

Methicillin-resistant Staphylococcus aureus (MRSA) causes life-threatening infections. Zinc oxide is well known as an effective antibacterial drug against many bacterial strains. We investigated the performance of zinc oxide nanorods synthesized by Albmiun as a biotemplate as an antibacterial drug in this study; the fabrication of zinc oxide nanorods was synthesized by sol-gel methods. We performed physicochemical characterization of zinc oxide nanorods by physiochemical techniques such as FTIR spectroscopy, X-ray diffraction, and TEM and investigation of their antimicrobial toxicity efficiency by MIC, ATPase activity assay, anti-biofilm activity, and kill time assays, as well as the mecA, mecR1, blaR1, blaZ, and biofilm genes (ica A, ica D, and fnb A) by using a quantitative RT-PCR assay and the penicillin-binding protein 2a (PBP2a) level of MRSA by using a Western blot. The data confirmed the fabrication of rod-shaped zinc oxide nanorods with a diameter in the range of 50 nm, which emphasized the formation of zinc oxide nanoparticles with regular shapes. The results show that zinc oxide nanorods inhibited methicillin-resistant S. aureus effectively. The MIC value was 23 µg/mL. The time kill of ZnO-NRs against MRSA was achieved after 2 h of incubation at 4MIC (92 µg/mL) and after 3 h of incubation at 2MIC (46 µg/mL), respectively. The lowest concentration of zinc oxide nanorods with over 75% biofilm killing in all strains tested was 32 µg/mL. Also, we examined the influence of the zinc oxide nanorods on MRSA by analyzing mecA, mecR1, blaR1, and blaZ by using a quantitative RT-PCR assay. The data obtained revealed that the presence of 2× MIC (46 µg/mL) of ZnO-NRs reduced the transcriptional levels of blaZ, blaR1, mecA, and mecR1 by 3.4-fold, 3.6-fold, 4-fold, and 3.8-fold, respectively. Furthermore, the gene expression of biofilm encoding genes (ica A, ica B, ica D, and fnb A) was tested using quantitative real-time reverse transcriptase-polymerase chain reaction (rt-PCR). The results showed that the presence of 2× MIC (46 µg/mL) of ZnO-NRs reduced the transcriptional levels of ica A, ica B, ica D, and fnb A. Also, the PBP2a level was markedly reduced after treatment with ZnO-NRs.

Fungicidal Activity of Zinc Oxide Nanoparticles against Azole-Resistant Aspergillus flavus Isolated from Yellow and White Maize.

The risk of resistance development and adverse effects on human health and the environment has increased in the last decade. Furthermore, many antifungal agents fail to inhibit the pathogenesis of azole-resistant Aspergillus flavus. In this report, we isolated and identified azole-resistant A. flavus isolates from two sources of maize (white and yellow maize). The susceptibilities of Aspergillus flavus isolates were investigated by conventional antifungals such as Terbinfine, Fluconazole, Ketoconazole, Voricazole, Amphotericin, and Nystatin. Then zinc oxide nanoparticles associated with Chlorella vulgaris, which are synthesized by using the precipitation method, were examined against isolated fungi. The results showed that twelve species of white corn were isolated out of fifty isolates, while the number of isolates from the yellow corn source was only four. Interestingly, the following antifungals have an impact effect against azole-resistant A. flavus isolates: the inhibition zones of ketoconazole, voricazole, and terbinafine were 40 mm, 20 mm, and 12 mm, respectively, while the remaining antifungal agents have no effect. Similarly, the inhibition zones of the following antifungal agents were as follows: 41 mm for Terbinfine, 13 mm for Voricazole, and 11 mm for Ketoconazole against Aspergillus flavus that was isolated from yellow corn. The physiochemical characterization of zinc oxide nanoparticles provides evidence that ZnO-NPs associate with Chlorella vulgaris and have been fabricated by the precipitation method with a diameter of 25 nm. The zinc oxide nanoparticle was then used to isolate azole-resistant A. flavus, and the results show that ZnO-NPs have an effect on azole-resistant A. flavus isolation. The inhibition zone of zinc oxide nanoparticles against A. flavus (that was isolated from white corn) was 50 mm with an MIC of 50 mg/mL, while the inhibition zone of zinc oxide nanoparticles against Azole-resistant A. flavus isolated from yellow corn was 14 nm with an MIC of 25 mg/mL, which indicated that zinc oxide nanoparticles gave a better result against Azole-resistant A. flavus isolated from maize.

Ultrastructural analysis of zinc oxide nanospheres enhances anti-tumor efficacy against Hepatoma.

Zinc oxide nanomaterial is a potential material in the field of cancer therapy. In this study, zinc oxide nanospheres (ZnO-NS) were synthesized by Sol-gel method using yeast extract as a non-toxic bio-template and investigated their physicochemical properties through various techniques such as FTIR, XR, DLS, and TEM. Furthermore, free zinc ions released from the zinc oxide nanosphere suspended medium were evaluated by using the ICP-AS technique. Therefore, the cytotoxicity of ZnO nanospheres and released Zn ions on both HuH7 and Vero cells was studied using the MTT assay. The data demonstrated that the effectiveness of ZnO nanospheres on HuH7 was better than free Zn ions. Similarly, ZnO-Ns were significantly more toxic to HuH7 cell lines than Vero cells in a concentration-dependent manner. The cell cycle of ZnO-Ns against Huh7 and Vero cell lines was arrested at G2/M. Also, the apoptosis assay using Annexin-V/PI showed that apoptosis of HuH7 and Vero cell lines by ZnO nanospheres was concentration and time-dependent. Caspase 3 assay results showed that the apoptosis mechanism may be intrinsic and extrinsic pathways. The mechanism of apoptosis was determined by applying the RT-PCR technique. The results revealed significantly up-regulated Bax, P53, and Cytochrome C, while the Bcl2 results displayed significant down-regulation and the western blot data confirmed the RT-PCR data. There is oxidative stress of the ZnO nanospheres and free Zn+2 ions. Results indicated that the ZnO nanospheres and free Zn+2 ions induced oxidative stress through increasing reactive oxygen species (ROS) and lipid peroxidation. The morphology of the HuH7 cell line after exposure to ZnO nanospheres at different time intervals revealed the presence of the chromatin condensation of the nuclear periphery fragmentation. Interestingly, the appearance of canonical ultrastructure features of apoptotic morphology of Huh7, Furthermore, many vacuoles existed in the cytoplasm, the majority of which were lipid droplets, which were like foamy cells. Also, there are vesicles intact with membranes that are recognized as swollen mitochondria.

Synergistic antibacterial activity of compact silver/magnetite core-shell nanoparticles core shell against Gram-negative foodborne pathogens.

The development of innovative antibacterial drugs against foodborne pathogens has led to an interest in novel materials such as nanomaterials. The unique features of nanomaterial qualify it for use as an antibacterial treatment. Noble metals and metal oxide nanoparticles, such as silver and magnetite nanoparticles, have been shown to be effective antibacterial medications against a range of microorganisms. In this work, Ag@Fe3O4 -NPs were fabricated by using a wet chemical reduction and modified co-precipitation techniques. The antibacterial efficiency of the Ag/Fe3O4 core shell nanoparticles was investigated by applying various techniques, such as the Kirby-Bauer Disk Diffusion test, minimum inhibitory concentration (MIC) and bactericidal concentration (MBC), Colony Forming Unit (CFU), and kill time assay. The toxicity mechanism of Ag@Fe3O4 -NPs against Salmonella typhimurium and Escherichia coli was studied by apoptosis and reactive oxygen species (ROS) assays. The data revealed that a cubic core was surrounded by a silver shell, which indicated the regular morphology of silver magnetite core shell nanoparticles without any aggregation. Furthermore, Ag@Fe3O4 -NPs is more toxic against S. typhimurium and E. coli than Ag-NPs and Fe3O4 NPs. The MIC values for Ag/Fe3O4 NPs against S. typhimurium and E. coli were 3.1 and 5.4 µg/ml, respectively, whereas the MIC values for Ag-NPs and MNPs against S. typhimurium and E. coli were 4.1 and 8.2 µg/ml for Ag-NPs and 6.9 and 10.3 µg/ml for MNPs. The results showed the ability of Ag@Fe3O4 -NPs to induce apoptosis by generating ROS. Also, the ability of Ag@Fe3O4 -NPs to liberate free Ag+ and generate ROS via the Haber-Weiss cycle may be a plausible mechanism to explain the toxicity of Ag@Fe3O4 -NPs - NPs.

Promoting Effect of Soluble Polysaccharides Extracted from Ulva spp. on Zea mays L. Growth.

Seaweeds can play a vital role in plant growth promotion. Two concentrations (5 and 10 mg/mL) of soluble polysaccharides extracted from the green macroalgae Ulva fasciata and Ulva lactuca were tested on Zea mays L. The carbohydrate and protein contents, and antioxidant activities (phenols, ascorbic, peroxidase, and catalase) were measured, as well as the protein banding patterns. The soluble polysaccharides at 5 mg/mL had the greatest effect on the base of all of the parameters. The highest effects of soluble polysaccharides on the Zea mays were 38.453, 96.76, 4, 835, 1.658, 7.462, and 38615.19, mg/mL for carbohydrates, proteins, phenol, µg ascorbic/mL, mg peroxidase/g dry tissue, and units/g tissue of catalase, respectively. The total number of protein bands (as determined by SDS PAGE) was not changed, but the density of the bands was correlated to the treatments. The highest band density and promoting effect were correlated to 5 mg/mL soluble polysaccharide treatments extracted from Ulva fasciata in Zea mays, which can be used as a biofertilizer.

Hearing screening in neonates with hyperbilirubinemia.

BACKGROUND: Hyperbilirubinemia is toxic to the auditory pathways and to the central nervous system, leaving sequelae such as hearing loss and encephalopathy. The damage to the auditory system occurs primarily within the brainstem and cranial nerve VIII, and manifests clinically as auditory neuropathy spectrum disorder. AIM: to establish the relationship that exists between hyperbilirubinemia at birth as a risk factor of neonatal hearing loss in children born in ABOU-EL-REISH hospital. MATERIALS AND METHODS: we carried out of 60 neonates categorized into two groups: Group (A n = 30), neonates with hyperbilirubinemia; Group (B n = 30), neonates without hyperbilirubinemia. RESULTS: Ten neonates were boys and twenty were girls from group A and eleven boys and nigh-teen girls from group B.There was significant statistical difference between group A and group B regarding ABR (P-value = 0.001) and a statistical difference regarding OAE (P-value = 0.103 in the right ear and 0.028 in the left ear). Also, our study shows the area under the curve and the diagnostic accuracy of total serum bilirubin (TSB) level for detection of hearing screening results (ABR) at a cut-off point 21 mg/dl (P-value = 0.008 and 0.009 in the right and left ears respectively.) CONCLUSIONS: Our study showed a relevant association between bilirubin levels and abnormal hearing screening results and the importance of combined screening with OAE and ABR tests.

Prevalence and molecular characterization of Shiga toxin-producing Escherichia coli isolates from human and sheep in Al-Madinah Al-Munawarah / Prevalencia y caracterización molecular de cepas de Escherichia coli productoras de toxina Shiga en seres humanos y ovejas de Al-Medina Al-Munawarah

Shiga toxin-producing Escherichia coli (STEC) strains have emerged as important foodborne pathogens of global public health concern, causing life-threatening diseases. Sheep and their products have been documented as important reservoirs for STECs, especially E. coli O157. The aim of this study was to investigate STECs from diarrheal human and sheep in Al-Madinah Al-Munawarah, Saudi Arabia. Fecal samples were collected between June and August, 2015 from diarrheal humans (n = 134) and sheep (n = 87). Presumptive E. coli human-and sheep-isolated strains were identified for their serotypes, the associated virulence genes (Shiga toxin [stx1 , stx2 ], haemolysin [ehxA] and intimin [eae]) by polymerase chain reaction and their susceptibility to antibiotics. Pulsed-field gel electrophoresis (PFGE) was used to demonstrate the genetic relatedness between Serotype O157:H7 human- and sheep-isolated strains. Forty eight (48/221; 21.7%) STECs were recovered from both human and sheep, their serotypes were as follows: O157:H7, O26:H11, O157:HNM, O26:HNM, O128:H2, O48:HNM, O111:HNM and OUT:HUT. Various virulence profiles and multiple antibiotic resistance were observed among the isolates. Twenty eight O157:H7 serotypes (17 human isolates and 11 sheep isolates) were identified in 13 PFGE pulsotypes, where human and sheep isolates were highly related. PFGE banding profiles together with serotypes and genotypes afford proof that human and sheep can be colonized and infected with similar E. coli O157:H7 strains. Our findings highlight the importance of epidemiological and microbiological surveillance of STECs; as well as the development of control measures to decrease risks associated with zoonotic O157:H7.

Las cepas de Escherichia coli (E. coli) productoras de toxina Shiga (STEC, del inglés Shiga toxin-producing E. coli) han surgido como importantes agentes patógenos de origen alimentario que son motivo de preocupación para la salud pública mundial, ya que provocan enfermedades potencialmente mortales. Se ha confirmado que las ovejas y sus productos son reservorios importantes para la STEC, especialmente E. coli O157. El objetivo de este estudio fue investigar STEC procedentes de deposiciones diarreicas humanas y ovinas en Al-Medina Al-Munawarah (Arabia Saudí). Se recogieron muestras fecales entre junio y agosto de 2015 de deposiciones diarreicas humanas (n = 134) y ovinas (n = 87). Se identificaron las presuntas cepas de E. coli humanas y ovinas por sus serotipos, los genes de virulencia asociados (toxina Shiga [stx1, stx2], hemolisina [ehxA] e intimina [eae]) por reacción en cadena de la polimerasa y la susceptibilidad a los antibióticos. Se utilizó la electroforesis en gel de campo pulsado (EGCP) para demostrar el parentesco genético entre el serotipo O157:H7 de las cepas humanas y el de las ovinas. Se aislaron 48 STEC (48/221; 21,7%) tanto humanas como ovinas y sus serotipos fueron los siguientes: O157:H7, O26:H11, O157:HNM, O26:HNM, O128:H2, O48:HNM, O111:HNM y OUT:HUT. Entre las cepas se observaron varios perfiles de virulencia y resistencia a múltiples antibióticos entre los aislamientos. Se identificaron 28 serotipos O157:H7 (17 cepas humanas y 11 cepas ovinas) en 13 pulsotipos de la EGCP, en los que las cepas humanas y ovinas estaban sumamente vinculadas. Los perfiles de bandeos de la EGCP, junto con los serotipos y genotipos, ofrecen una prueba de que seres humanos y ovejas pueden ser colonizados e infectados por cepas similares de E. coli O157:H7. Nuestros resultados destacan la importancia de la vigilancia epidemiológica y microbiológica de STEC, así como del desarrollo de medidas de control para reducir los riesgos asociados con la O157:H7 zoonótica.

A novel mutation in exon 1 of GATA4 in Egyptian patients with congenital heart disease.

BACKGROUND/AIM: Congenital heart disease (CHD) is a common birth defect. Many studies have reported GATA4 mutations in patients with CHD, mainly septal defects. This study aimed to investigate the GATA4 exon 1 mutation in Egyptian patients with isolated congenital heart defects as a possible causative mutation. MATERIALS AND METHODS: Screening for mutations or any sequence variations in exon 1 of the GATA4 gene was carried out by PCR amplification followed by direct sequencings in 165 Egyptian patients with different nonsyndromic congenital heart diseases and 93 controls who were matched in terms of age and sex. Thorough clinical assessments were done for all subjects, along with X-ray, 2D echocardiography, and Doppler examinations. RESULTS: The most common CHD among our cases was isolated ventricular septal defect (VSD) in 47.3% (78/165), followed by isolated atrial septal defect. A novel nonsynonymous sequence variation in fragment 2 (P193H) of exon 1 of GATA4 was detected in 15 (9.1%) of the subjects with septal defects. This mutation was not seen in any of the control group subjects. CONCLUSION: There is a high prevalence of exon 1 GATA4 mutation (9.1%) in our study compared to other studies in different populations, which may correlate with different ethnic populations.

Trafficking of methicillin-resistant staphylococci and co-colonization with vancomycin-resistant enterococci.

OBJECTIVES: To determine the trafficking of methicillin-resistant staphylococci between the hospital and community as well as the occurrence of co-colonization with vancomycin-resistant enterococci (VRE). SUBJECTS AND METHODS: From November 2005 to April 2006, methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-resistant coagulase-negative Staphylococcus (MRCoNS)-positive patients at the Salmaniya Medical Complex, Bahrain were assessed for VRE co-colonization. Characterization of vancomycin resistance genotype by PCR was carried out. Close family contacts were screened for MRSA and pulsed-field gel electrophoresis (PFGE) analysis of MRSA isolates from patient-family member pairs was conducted. RESULTS: One hundred and eighty-two patients (93 MRSA; 89 MRCoNS) and 356 family members were enrolled. Seven MRSA and 41 MRCoNS strains were isolated from the family members. PFGE analysis revealed the presence of variants of a single MRSA clone among patients and their relatives. A total of 112 patients (62 MRSA; 50 MRCoNS) provided stool for VRE screening. Of these 13 stool specimens (11.6%) were VRE-positive. All the VRE isolates were from MRSA-positive patients, thus positivity rate among MRSA patients was 20.9% (n/N = 13/62). These were predominantly Enterococcus gallinarum with vanC1 genotype and one strain was Enterococcus faecium (vanB genotype). Two E. gallinarum isolates harbored an additional vanB gene. The majority of VRE isolates were from patients in medical and surgical units (n/N = 10/13; 77%). Male gender, prolonged hospitalization and presence of co-morbidities were significantly associated with MRSA/VRE co-colonization (p < 0.05). CONCLUSION: MRSA/VRE co-colonization with MRSA trafficking between the hospital and community environment is a public health concern occurring in our setting.

Keratinolytic activity of purified alkaline keratinase produced by Scopulariopsis brevicaulis (Sacc.) and its amino acids profile.

Sodium dodecyl sulfate-polyacrlyamide gel electrophoresis (SDS-PAGE) was used to assess the purity and molecular weight of the previously purified alkaline keratinase enzyme of Scopulariopsis brevicaulis. The enzyme was homogenous, as seen by a single band of protein, and had an apparent molecular weight of 28.5 kDa. Amino acid profile of the purified keratinase revealed that it was composed of 14 different amino acids with high proportions of glutamic acid (20.86%), alanine (14.52%), glycine (14.21%), leucine (8.59%) and serine (7.81%). The enzyme contained moderate amounts of valine (6.01%), threonine (5.58%) and phenyl alanine (5.22%). The purified enzyme of S. brevicaulis exerted a potent keratinolytic activity and was capable to hydrolyze different keratinaceous materials with highest activity on chicken feathers followed by human nails and human hair.

Pattern of antibiotic susceptibility in Campylobacter jejuni isolates of human and poultry origin.

Campylobacter jejuni antibiotic resistance is rising with a variable geographical pattern; but there is limited data from the Arabian Gulf region. We assessed the sensitivity of human (117) and chicken (33) C. jejuni isolates to erythromycin, ciprofloxacin, tetracycline and trimethoprim-sulfamethoxazole by agar dilution, disc diffusion and the E test. Only 2 human isolates were resistant to erythromycin. In contrast, over 80% of chicken and human isolates were resistant to ciprofloxacin. A significantly higher proportion of chicken isolates than human isolates were resistant to tetracycline, with much higher MIC(50) values (P < 0.001). The MIC(90) for trimethoprim-sulfamethoxazole by agar dilution was 40 microg/ml. Comparison of the results of the agar dilution method and E test showed 1 major disagreement and 8 minor disagreements for erythromycin, 4 major disagreements for ciprofloxacin and 23 disagreements for tetracycline (19 were major disagreements). This was the first study to describe the pattern of antibiotic resistance in Campylobacter isolates in this region; the results indicate a high degree of erythromycin sensitivity that validates the continued use of this agent as a first-line therapy for Campylobacter enteritis. These findings have wide implications because of the large, highly mobile expatriate population in this setting. In addition, the correlation between agar dilution and disc diffusion supports the use of the latter as an alternative susceptibility testing method for Campylobacter.

A potent chitinolytic activity of Alternaria alternata isolated from Egyptian black sand.

Eight fungal species characterized by chitinolytic activity were isolated from Egyptian black sand collected from Rosetta coast. Genus Aspergillus and Alternaria alternata exhibited the highest density (> 40% of the total count, each) on the isolation plates containing different treatments of native shrimp shell chitin. Genus Aspergillus was represented by A. flavus, A. niger, A. foetidus and A. tungius, with the former species being the most dominant. The other species were Cladosporium herbarum, Fusarium equisitum (5.71% of the total count, each) and Dendryphiella vinosa (3.21% of the total count). The isolated species were screened for chitinase production on agar plates containing 0.2% colloidal chitin. The chitinolytic activity of each individual was not always correlated with its density on the isolation plates. Alternaria alternata was the most promising species for chitinase excretion. The use of colloidal chitin (1.5%) as a sole carbon source was superior for the enzyme production by A. alternata. Maximum enzyme yield was obtained after 7 days incubation at 30 degrees C with shaking (150 rev min(-1)), with an initial pH value of the growth medium at 5.0. Presence of NaNO3 (0.3%), the best nitrogen source, and CaCl2 (100 microg/ml) stimulated the induction of the enzyme. The crude A. alternata chitinase revealed a potential insecticidal effect on the larvae of fruitfly (82% mortality) and could degrade crude shrimp shell waste.

Induced resistance in tomato plants by IAA against Fusarium oxysporum lycopersici.

The phytohormone IAA (indol-3-acetic acid) was tested in vitro on growth of tomato wilt pathogen Fusarium oxysporum lycopersici. The hormone reduced spore germination, mycelial dry weight and protein content. Such reduction was matched with the elevation in the hormone concentration. The in vivo application of IAA to soil of the uninoculated plants (controls) improved growth and yielded longer shoot and root, particularly at low concentrations. Moreover, the hormone could prevent completely any chance for disease incidence by soil pathogens. Presence of IAA in soil of inoculated plants not only reduced the infection rate but also increased plant growth, causing that they appeared healthy and normal. Disease suppression in tomato plants, exerted by application of IAA, was achieved through either increasing plant growth, exerting a direct harmful effect on the target pathogen and/or inducing resistance in host tissue. The induced resistance was correlated with induction of certain secondary metabolites which may have a role in increasing tolerance in tomato plants to the pathogen.

Impact of industrial waste water effluents on mycoflora of the shore sediments of the 3rd oxidation pond, with reference to biosorption of heavy metals.

The third oxidation pond at 10th of Ramadan desert receives a number of industrial waste water effluents contaminated with the heavy metal ions Zn, Cd, Cu and Ni. The species diversity and fungal community structure of seven different sites at the onshore sediments and offshore were studied. Mycological analysis resulted in isolation of 3912 fungal colonies, 11.7% of this count were recovered from the onshore sediment sites (4 sites) whereas 88.3% were from the offshore sites (3 sites), in the desert. Fungal counts and species diversity at the onshore sites tend to increase with increasing distance far from the waste water input. A complete accordance was observed among the total fungal counts and species variabilities with organic matter content at each sampling site. This relationship was reversed in case of heavy metal contents with both counts and diversity. Seventeen fungal species belonging to seven genera were isolated from all sites under study. Aspergillus spp. constituted the majority of the isolates (51.7% of the total isolates), followed by Curvularia, Cephalosporium, and Humicola. Of nine isolated Aspergillus spp., A. humicola was the most dominant (37.4% of the total catch) and appeared at all polluted sites. Therefore, A. humicola was chosen to investigate its potential for heavy metals sorption from the contaminated waste water effluent. Four days old biomass pellets could sorb a large amount of heavy metals according to the following sequence: Zn>Cd>Cu>Ni ions. Agitation significantly increased Zn and Cd sorption, but not Cu and Ni. Heavy metals sorption took place at a wide pH range and particularly increased at alkaline pH (8-9).