Search results for: bacterial ring rot

Authors: Mrinal Gupta, Mohammad Rumman, Babita Singh Abbas Ali Mahdi, Shivani Pandey

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Introduction: Berberine (BBR), a bioactive compound isolated from Coptidis Rhizoma, possesses diverse pharmacological activities, including anti-bacterial, anti-inflammatory, antitumor, hypolipidemic, and anti-diabetic. However, its role as an anti-diabetic agent in animal models of dexamethasone (Dex)-induced diabetes remains unknown. Studies have shown that natural compounds, including aloe, caper, cinnamon, cocoa, green and black tea, and turmeric, can be used for treating Type 2 diabetes mellitus (DM). Compared to conventional drugs, natural compounds have fewer side effects and are easily available. Herein, we studied the anti-diabetic effects of BBR in a mice model of Dex-induced diabetes. Methods: HepG2 cell line was used for glucose release and glycogen synthesis studies. Cell proliferation was measured by methylthiotetrazole (MTT) assay. For animal studies, mice were treated with Dex (2 mg/kg, i.m.) for 30 days and the effect of BBR at the doses 100, 200, and 500 mg/kg (p.o.) was analyzed. Glucose, insulin, and pyruvate tests were performed to evaluate the development of the diabetic model. An echo MRI was performed to assess the fat mass. Further, to elucidate the mechanism of action of BBR, mRNA expression of genes regulating gluconeogenesis, glucose uptake, and glycolysis were analyzed. Results: In vitro BBR had no impact on cell viability up to a concentration of 50μM. Moreover, BBR suppressed the hepatic glucose release and improved glucose tolerance in HepG2 cells. In vivo, BBR improved glucose homeostasis in diabetic mice, as evidenced by enhanced glucose clearance, increased glycolysis, elevated glucose uptake, and decreased gluconeogenesis. Further, Dex treatment increased the total fat mass in mice, which was ameliorated by BBR treatment. Conclusion: BBR improves glucose tolerance by increasing glucose clearance, inhibiting hepatic glucose release, and decreasing obesity. Thus, BBR may become a potential therapeutic agent for treating glucocorticoid-induced diabetes and obesity in the future.

Keywords: glucocorticoid, hyperglycemia, berberine, HepG2 cells, insulin resistance, glucose

Procedia PDF Downloads 64

Authors: Andrew Cummings, Rhianne Boag, Sarah Bryson, Gordon Turner

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Packages designed for transport of radioactive material must satisfy rigorous safety regulations specified by the International Atomic Energy Agency (IAEA). Higher Activity Waste (HAW) transport packages have to maintain containment of their contents during normal and accident conditions of transport (NCT and ACT). To ensure containment criteria is satisfied these packages are required to be leak-tight in all transport conditions to meet allowable activity release rates. Package design safety reports are the safety cases that provide the claims, evidence and arguments to demonstrate that packages meet the regulations and once approved by the competent authority (in the UK this is the Office for Nuclear Regulation) a licence to transport radioactive material is issued for the package(s). The standard approach to demonstrating containment in the RWM transport safety case is set out in BS EN ISO 12807. In this document a method for measuring a leak rate from the package is explained by way of a small interspace test volume situated between two O-ring seals on the underside of the package lid. The interspace volume is pressurised and a pressure drop measured. A small interspace test volume makes the method more sensitive enabling the measurement of smaller leak rates. By ascertaining the activity of the contents, identifying a releasable fraction of material and by treating that fraction of material as a gas, allowable leak rates for NCT and ACT are calculated. The adherence to basic safety principles in ISO12807 is very pessimistic and current practice in the demonstration of transport safety, which is accepted by the UK regulator. It is UK government policy that management of HAW will be through geological disposal. It is proposed that the intermediate level waste be transported to the geological disposal facility (GDF) in large cuboid packages. This poses a challenge for containment demonstration because such packages will have long seals and therefore large interspace test volumes. There is also uncertainty on the releasable fraction of material within the package ullage space. This is because the waste may be in many different forms which makes it difficult to define the fraction of material released by the waste package. Additionally because of the large interspace test volume, measuring the calculated leak rates may not be achievable. For this reason a justification for a lower releasable fraction of material is sought. This paper considers the use of aerosol processes to reduce the releasable fraction for both NCT and ACT. It reviews the basic coagulation and removal processes and applies the dynamic aerosol balance equation. The proposed solution includes only the most well understood physical processes namely; Brownian coagulation and gravitational settling. Other processes have been eliminated either on the basis that they would serve to reduce the release to the environment further (pessimistically in keeping with the essence of nuclear transport safety cases) or that they are not credible in the conditions of transport considered.

Keywords: aerosol processes, Brownian coagulation, gravitational settling, transport regulations

Procedia PDF Downloads 117

Authors: Satwik Majumder, Dongyun Jung, Jennifer Ronholm, Saji George

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Bovine mastitis is the most common infectious disease in dairy cattle, with major economic implications for the dairy industry worldwide. Continuous monitoring for the emergence of antimicrobial resistance (AMR) among bacterial isolates from dairy farms is vital not only for animal husbandry but also for public health. In this study, the prevalence of AMR in 113 Escherichia coli isolates from cases of bovine clinical mastitis in Canada was investigated. Kirby-Bauer disk diffusion test with 18 antibiotics and microdilution method with three heavy metals (copper, zinc, and silver) was performed to determine the antibiotic and heavy-metal susceptibility. Resistant strains were assessed for efflux and ß-lactamase activities besides assessing biofilm formation and hemolysis. Whole-genome sequences for each of the isolates were examined to detect the presence of genes corresponding to the observed AMR and virulence factors. Phenotypic analysis revealed that 32 isolates were resistant to one or more antibiotics, and 107 showed resistance against at least one heavy metal. Quinolones and silver were the most efficient against the tested isolates. Among the AMR isolates, AcrAB-TolC efflux activity and ß-lactamase enzyme activities were detected in 13 and 14 isolates, respectively. All isolates produced biofilm but with different capacities, and 33 isolates showed α-hemolysin activity. A positive correlation (Pearson r = +0.89) between efflux pump activity and quantity of biofilm was observed. Genes associated with aggregation, adhesion, cyclic di-GMP, quorum sensing were detected in the AMR isolates, corroborating phenotype observations. This investigation showed the prevalence of AMR in E. coli isolates from bovine clinical mastitis. The results also suggest the inadequacy of antimicrobials with a single mode of action to curtail AMR bacteria with multiple mechanisms of resistance and virulence factors. Therefore, it calls for combinatorial therapy for the effective management of AMR infections in dairy farms and combats its potential transmission to the food supply chain through milk and dairy products.

Keywords: antimicrobial resistance, E. coli, bovine mastitis, antibiotics, heavy-metals, efflux pump, ß-lactamase enzyme, biofilm, whole-genome sequencing

Procedia PDF Downloads 216

Authors: Himalaya Patir, Bitupon Baruah, Sanjay Gayary, Subhajit Ray

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In recent age significant importance is given for the development of nutritious and health beneficial foods. Fruit juices collected from different fruits when blended that improves not only the physicochemical and nutritional properties but also enhance the sensorial or organoleptic properties. The study was carried out to determine the physico-chemical, nutritional, microbiological analysis and sensory evaluation of mixed fruit juice blend. Juice of orange (Citrus sinensis), apple (Malus domestica), mosambi (Citrus limetta) were blended in the ratio of sample-I (30% apple:30% orange:40% mosambi), sample-II ( 40% apple :30% orange :30% mosambi), sample-III (30% apple :40% orange :30% mosambi) , sample-IV (50% apple :30% orange :20% mosambi), sample-V (30% apple:20% orange:50% mosambi), sample-VI (20% apple :50% orange :30% mosambi) to evaluate all quality characteristics. Their colour characteristics in terms of hue angle, chroma and colour difference (∆E) were evaluated. The physico-chemical parameters analysis carried out were total soluble solids (TSS), total titratable acidity (TTA), pH, acidity (FA), volatile acidity (VA), pH, and vitamin C. There were significant differences (p˂0.05) in the TSS of the samples. However, sample-V (30% apple: 20% orange: 50% mosambi) provides the highest TSS of 9.02gm and significantly differed from other samples (p˂0.05). Sample-IV (50% apple: 30% orange: 20% mosambi) was shown the highest titratable acidity (.59%) in comparison to other samples. The highest value of pH was found as 5.01 for sample-IV (50% apple: 30% orange: 20% mosambi). Sample-VI (20% apple: 50% orange :30% mosambi) blend has the highest hue angle, chroma and colour changes of 72.14,25.29 and 54.48 and vitamin C, i.e. Ascorbic acid (.33g/l) content compared to other samples. The nutritional compositions study showed that, sample- VI (20% apple: 50% orange: 30% mosambi) has the significantly higher carbohydrate (51.67%), protein (.78%) and ash (1.24%) than other samples, while sample-V (30% apple: 20% orange: 50% mosambi) has higher dietary fibre (12.84%) and fat (2.82%) content. Microbiological analysis of all samples in terms of total plate count (TPC) ranges from 44-60 in 101 dilution and 4-5 in 107 dilutions and was found satisfactory. Moreover, other pathogenic bacterial count was found nil. The general acceptability of the mixed fruit juice blend samples were moderately liked by the panellists, and sensorial quality studies showed that sample-V (30% apple: 20% orange: 50% mosambi) contains highest overall acceptability of 8.37 over other samples and can be considered good for consumption.

Keywords: microbiological, nutritional, physico-chemical, sensory properties

Procedia PDF Downloads 178

Authors: Ade O. Oyewole, Sunday O. Okoh, Ruth O. Ishola, Adenike D. Odusote, Chima C. Igwe, Gloria N. Elemo, Anthony I. Okoh

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Azadirachta indica (Neem tree) also referred to as an all-purpose tree is used in a wide range of medical preparations in tropical and subtropical countries for prevention and management of various livestock, crops products and human diseases. In Nigeria however, the potentials of this plant have not been fully exploited thus it causes an environmental nuisance during the fruiting season. With a rise in the demand for herbal personal care products globally extracts from different parts of the neem plant were used as the bio-active ingredients in the formulation of personal care products. In this study, formulated neem soap, body cream, lotion, toothpaste and shampoo are analyzed to determine their antibacterial, antifungal, and toxicity properties. The efficacies of these products for management of infectious diseases, both oral and dermal, were also investigated in vitro. Oil from the neem seeds obtained using a mechanical press and acetone extracts of both the neem bark and leaves obtained by the maceration method were used in the formulation and production of the neem personal care products. The antimicrobial and toxicity properties of these products were investigated by agar diffusion, and haemolytic methods respectively. The five neem products (NPs) exhibited strong antibacterial activities against four multi–drug resistant pathogenic and three none pathogenic bacterial strains (Escherichia coli (180), Listeria ivanovii, Staphylococcus aureus, Enterobacter cloacae, Vibro spp., Streptococcus uberis, Mycobacterium smegmatis), except the neem lotion with insignificant activity against E. coli and S. aureus. The minimum inhibitory concentration (MIC) range was between 0.20-0.40 mg/ mL. The 5 NPs demonstrated moderate activity against three clinical dermatophytes isolates (Tinea corporis, Tinea capitis, and Tinea cruiz) as well as one fungal strain (Candida albican) with the MIC ranging between 0.30 - 0.50 mg/ mL and 0.550 mg/mL respectively. The soap and shampoo were the most active against test bacteria and fungi. The haemolytic analysis results on the 5 NPs indicated none toxicity at 0.50 mg/ mL in sheep red blood cells (SRBC).

Keywords: antimicrobial, Azadirachta indica, multi–drug resistant pathogenic bacteria, personal care products

Procedia PDF Downloads 270

Authors: Portia Murphy, Danica Vasic, Anoja W. Gunaratne, Encarnita Sitchon, Teresita Tugonon, Marou Ison, Antoinette Le Busque, Christelle Pagonis, Thomas J. Borody

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Irritable bowel syndrome (IBS) is a chronic gastrointestinal disorder that affects an estimated 11% of the population globally with the most predominant symptoms being abdominal pain, bloating and altered bowel movements. All age groups suffer from IBS although the prevalence of IBS decreases for age groups over 50 years. Women are more likely to suffer from IBS than men. IBS can be categorized into 3 groups based on the type of altered bowel movement: diarrhea-predominant IBS (IBS-D), constipation-predominant IBS (IBS-C) and IBS with mixed bowel habit (IBS-M). The contribution of the gut microbiome to the etiology of IBS is becoming increasingly recognized with rising use of anti-microbial agents. Previous studies on vancomycin and rifaximin used as monotherapy or in combination have been conducted mainly on IBS-C and showed marked improvements in the symptoms. According to our knowledge, no studies reported using these two combinations of antibiotics for IBS-D. Here, we report a consecutive cohort of 18 patients treated with both vancomycin and rifaximin for IBS-D. These patients’ records were reviewed retrospectively. In this cohort, patients ages were between 24-74 years (mean 44 years) and 9 were female. Baseline all patients had diarrhea, 4 with mucus and one with blood. Patients reported other symptoms were abdominal pain (n=11) bloating (n=9), flatulence (n=7), fatigue (n=4) and nausea (n=3). Patients treatments were personalized according to their symptom severity and tolerability and were treated with combination of rifaximin (500 - 3000mg/d) and vancomycin (500mg - 1500mg/d) for an ongoing period. Follow-ups were conducted between 2-32 weeks’ time. Of all patients, 89% patients reported improvement of the symptoms, 1 reported no change and 1 patient’s symptoms got worse. The mechanism of action for both vancomycin and rifaximin involves the inhibition of bacterial cell wall and protein synthesis respectively. The role of these medications in improving the symptoms of this cohort suggests that IBS-D may be microbiome infection driven. In this cohort, similar patient presentations to Clostridium difficile, as well as symptom improvement with the use of rifaximin and particularly vancomycin, suggest that the infectious agent may be an unidentified Clostridium. These preliminary results offer an alternative etiology for IBS-D not previously considered and open the avenue for new research.

Keywords: clostridium deficile, diarrhea predominant Irritable Bowel Syndrome, microbiome, vancomycin/rifaximin combination

Procedia PDF Downloads 130

Authors: Swati Tomar, Sunil Kumar Gupta

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Anammox is a promising and cost effective alternative to conventional treatment systems that facilitates direct oxidation of ammonium nitrogen under anaerobic conditions with nitrite as an electron acceptor without addition of any external carbon sources. The present study investigates the process kinetics of laboratory scale anammox hybrid reactor (AHR) which combines the dual advantages of attached and suspended growth. The performance & behaviour of AHR was studied under varying hydraulic retention time (HRTs) and nitrogen loading rate (NLRs). The experimental unit consisted of 4 numbers of 5L capacity anammox hybrid reactor inoculated with mixed seed culture containing anoxic and activated sludge. Pseudo steady state (PSS) ammonium and nitrite removal efficiencies of 90.6% and 95.6%, respectively, were achieved during acclimation phase. After establishment of PSS, the performance of AHR was monitored at seven different HRTs of 3.0, 2.5, 2.0, 1.5, 1.0, 0.5 and 0.25 d with increasing NLR from 0.4 to 4.8 kg N/m3d. The results showed that with increase in NLR and decrease in HRT (3.0 to 0.25 d), AHR registered appreciable decline in nitrogen removal efficiency from 92.9% to 67.4 %, respectively. The HRT of 2.0 d was considered optimal to achieve substantial nitrogen removal of 89%, because on further decrease in HRT below 1.5 days, remarkable decline in the values of nitrogen removal efficiency were observed. Analysis of data indicated that attached growth system contributes an additional 15.4 % ammonium removal and reduced the sludge washout rate (additional 29% reduction). This enhanced performance may be attributed to 25% increase in sludge retention time due to the attached growth media. Three kinetic models, namely, first order, Monod and Modified Stover-Kincannon model were applied to assess the substrate removal kinetics of nitrogen removal in AHR. Validation of the models were carried out by comparing experimental set of data with the predicted values obtained from the respective models. For substrate removal kinetics, model validation revealed that Modified Stover-Kincannon is most precise (R2=0.943) and can be suitably applied to predict the kinetics of nitrogen removal in AHR. Lawrence and McCarty model described the kinetics of bacterial growth. The predicted value of yield coefficient and decay constant were in line with the experimentally observed values.

Keywords: anammox, kinetics, modelling, nitrogen removal, sludge wash out rate, AHR

Procedia PDF Downloads 316

Authors: Michelle Themalil, Celia Bueno, Rulla Al- Araji

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Objective and Study: There are no established guidelines for antibiotic prophylaxis in children with central venous catheters (CVCs) on home parenteral nutrition (HPN), leading to varying practices across UK Centres. We hypothesize that children with intestinal failure are at increased risk for bacteraemia due to altered anatomy, dysmotility, inflammation, biofilm formation in long-term CVCs, and the use of central lines during procedures. Given the bacteraemia rates of up to 8% in upper and 25% in lower endoscopy for adults without central lines, we argue that prophylactic antibiotics are reasonable, given the increased risks faced by this high-risk group of children. Methods: We conducted a five-year review of patients with central venous access receiving home parenteral nutrition (HPN) who underwent endoscopies with antibiotic prophylaxis at our center (tertiary). We documented and analyzed post-procedure infections and their associated risk factors. Results: A total of 15 patients on HPN underwent 29 endoscopic procedures, including 4 upper, 9 combined upper and lower, and 16 combined upper, lower, and ileoscopy. Confirmed infection rates remained at 0% up to 28 days post-procedure. The agreed-upon prophylaxis regimen was implemented, with ciprofloxacin and metronidazole administered as the primary antibiotics. Notably, only 51.7% of patients received a peripheral cannula despite recommendations to avoid central line use during anesthesia, and 20.6% had small intestinal bacterial overgrowth. Conclusions: This study is the first to investigate post-endoscopy infection rates in pediatric patients on HPN. Despite a small sample size, we observed a 0% infection rate, significantly lower than reported rates in adults. These findings suggest that further research is warranted to explore the implications of antibiotic prophylaxis in this unique patient cohort and to establish guidelines that may enhance patient safety during endoscopic procedures.

Keywords: post endosopy infections, central venous access, home parenteral nutrition, intestinal failure

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Authors: Arturo Galván, Jatziri Y. Moreno-Martínez, Israel Enrique Herrera Díaz, José Ramón Gasca Tirado

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In recent years, one of the methods most used for the construction of tunnels in soft soil is the shield-driven tunneling. The advantage of this construction technique is that it allows excavating the tunnel while at the same time a primary lining is placed, which consists of precast segments. There are joints between segments, also called longitudinal joints, and joints between rings (called as circumferential joints). This is the reason because of this type of constructions cannot be considered as a continuous structure. The effect of these joints influences in the rigidity of the segmental lining and therefore in its structural response. A parametric study was performed to take into account the effect of different parameters in the structural response of typical segmental tunnels built in soft soil by using non-linear numerical models based on Finite Element Method by means of the software package ANSYS v. 11.0. In the first part of this study, two types of numerical models were performed. In the first one, the segments were modeled by using beam elements based on Timoshenko beam theory whilst the segment joints were modeled by using inelastic rotational springs considering the constitutive moment-rotation relation proposed by Gladwell. In this way, the mechanical behavior of longitudinal joints was simulated. On the other hand for simulating the mechanical behavior of circumferential joints elastic springs were considered. As well as, the stability given by the soil was modeled by means of elastic-linear springs. In the second type of models, the segments were modeled by means of three-dimensional solid elements and the joints with contact elements. In these models, the zone of the joints is modeled as a discontinuous (increasing the computational effort) therefore a discrete model is obtained. With these contact elements the mechanical behavior of joints is simulated considering that when the joint is closed, there is transmission of compressive and shear stresses but not of tensile stresses and when the joint is opened, there is no transmission of stresses. This type of models can detect changes in the geometry because of the relative movement of the elements that form the joints. A comparison between the numerical results with two types of models was carried out. In this way, the hypothesis considered in the simplified models were validated. In addition, the numerical models were calibrated with (Lab-based) experimental results obtained from the literature of a typical tunnel built in Europe. In the second part of this work, a parametric study was performed by using the simplified models due to less used computational effort compared to complex models. In the parametric study, the effect of material properties, the geometry of the tunnel, the arrangement of the longitudinal joints and the coupling of the rings were studied. Finally, it was concluded that the mechanical behavior of segment and ring joints and the arrangement of the segment joints affect the global behavior of the lining. As well as, the effect of the coupling between rings modifies the structural capacity of the lining.

Keywords: numerical models, parametric study, segmental tunnels, structural response

Procedia PDF Downloads 229

Authors: Estelle Arbellay, Lori D. Daniels, Shawn D. Mansfield, Alice S. Chang

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Mountain pine beetle (MPB) outbreaks are currently devastating lodgepole pine forests in western North America, which are also widely disturbed by frequent wildfires. Both MPB and fire can leave scars on lodgepole pine trees, thereby diminishing their commercial value and possibly compromising their utilization in solid wood products. In order to fully exploit the affected resource, it is crucial to understand how wounding from these two disturbance agents impact wood properties. Moreover, previous research on lodgepole pine has focused solely on sound wood and stained wood resulting from the MPB-transmitted blue fungi. By means of a quantitative multi-proxy approach, we tested the hypotheses that (i) wounding (of either MPB or fire origin) caused significant changes in wood properties of lodgepole pine and that (ii) MPB-induced wound effects could differ from those induced by fire in type and magnitude. Pith-to-bark strips were extracted from 30 MPB scars and 30 fire scars. Strips were cut immediately adjacent to the wound margin and encompassed 12 rings from normal wood formed prior to wounding and 12 rings from wound wood formed after wounding. Wood properties evaluated within this 24-year window included ring width, relative wood density, cellulose crystallinity, fibre dimensions, and carbon and nitrogen concentrations. Methods used to measure these proxies at a (sub-)annual resolution included X-ray densitometry, X-ray diffraction, fibre quality analysis, and elemental analysis. Results showed a substantial growth release in wound wood compared to normal wood, as both earlywood and latewood width increased over a decade following wounding. Wound wood was also shown to have a significantly different latewood density than normal wood 4 years after wounding. Latewood density decreased in MPB scars while the opposite was true in fire scars. By contrast, earlywood density was presented only minor variations following wounding. Cellulose crystallinity decreased in wound wood compared to normal wood, being especially diminished in MPB scars the first year after wounding. Fibre dimensions also decreased following wounding. However, carbon and nitrogen concentrations did not substantially differ between wound wood and normal wood. Nevertheless, insect-feeding and fire-heating wounding were shown to significantly alter most wood properties of lodgepole pine, as demonstrated by the existence of several morphological anomalies in wound wood. MPB and fire generally elicited similar anomalies, with the major exception of latewood density. In addition to providing quantitative criteria for differentiating between biotic (MPB) and abiotic (fire) disturbances, this study provides the wood industry with fundamental information on the physiological response of lodgepole pine to wounding in order to evaluate the utilization of scarred trees in solid wood products.

Keywords: elemental analysis, fibre quality analysis, lodgepole pine, wood properties, wounding, X-ray densitometry, X-ray diffraction

Procedia PDF Downloads 319

Authors: Simona Dostalova, Dita Munzova, Ana Maria Jimenez Jimenez, Marketa Vaculovicova, Vojtech Adam, Rene Kizek

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The boom of nanomedicine in recent years has led to the development of numerous new nanomaterials that can be used as nanocarriers in the drug delivery. These nanocarriers can either be synthetic or natural-based. The disadvantage of many synthetic nanocarriers is their toxicity in patient’s body. Protein cages that can naturally be found in human body do not exhibit such disadvantage. However, the release of cargo from some protein cages in target cells can be problematic. As a special type of protein cages can serve the capsid of many viruses, including phage. Phages infect bacterial cells; therefore they are not harmful to human cells. The targeting of phage particles to cancer cells can be solved by producing of empty phage capsids during which the targeting moieties (e.g. peptides) can be cloned into genes of phage capsid to decorate its surface. Moreover, the produced capsids do not contain viral nucleic acid and are therefore not infectious to beneficial bacteria in the patient’s body. The protein cage composed of viral capsid is larger than other frequently used apoferritin cage but its size is still small enough to benefit from passive targeting by Enhanced Permeability and Retention effect. In this work, bacteriophage λ was used, both whole and its empty capsid for delivery of different cytotoxic drugs (cisplatin, carboplatin, oxaliplatin, etoposide and doxorubicin). Large quantities of phage λ were obtained from phage λ-producing strain of E. coli cultivated in medium with 0.2 % maltose. After killing of E. coli with chloroform and its removal by centrifugation, the phage was concentrated by ultracentrifugation at 130 000 g and 4 °C for 3 h. The encapsulation of the drugs was performed by infusion method and four different concentrations of the drugs were encapsulated (200; 100; 50; 25 µg/ml). Free molecules of drugs were removed by dialysis. The encapsulation was verified using spectrophotometric and electrochemical methods. The amount of encapsulated drug linearly increased with the amount of applied drug (determination coefficient R2=0.8013). 76% of applied drug was encapsulated in phage λ particles (concentration of 10 µg/ml), even with the highest applied concentration of drugs, 200 µg/ml. Only 1% of encapsulated drug was detected in phage DNA. Similar results were obtained with encapsulation in phage empty capsid. Therefore, it can be concluded that the encapsulation of drugs into phage particles is efficient and mostly occurs by interaction of drugs with protein capsid.

Keywords: cytostatics, drug delivery, nanocarriers, phage capsid

Procedia PDF Downloads 493

Authors: Ane Escobar, Paula Angelomé, Mihaela Delcea, Marek Grzelczak, Sergio Enrique Moya

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The antibacterial capacity of bone-anchoring implants can be improved by the use of antibiotics that can be delivered to the media after the surgery. Mesoporous films have shown great potential in drug delivery for orthopedic applications, since pore size and thickness can be tuned to produce different surface area and free volume inside the material. This work shows the synthesis of mesoporous titania films (MTF) by sol-gel chemistry and evaporation-induced self-assembly (EISA) on top of glass substrates. Pores with a diameter of 12nm were observed by Transmission Electron Microscopy (TEM). A film thickness of 100 nm was measured by Scanning Electron Microscopy (SEM). Gentamicin was used to study the antibiotic delivery from the film by means of High-performance liquid chromatography (HPLC). The Staphilococcus aureus strand was used to evaluate the effectiveness of the penicillin loaded films toward inhibiting bacterial colonization. MC3T3-E1 pre-osteoblast cell proliferation experiments proved that MTFs have a good biocompatibility and are a suitable surface for MC3T3-E1 cell proliferation. Moreover, images taken by Confocal Fluorescence Microscopy using labeled vinculin, showed good adhesion of the MC3T3-E1 cells to the MTFs, as well as complex actin filaments arrangement. In order to improve cell proliferation Bone Morphogenetic Protein-2 (BMP-2) was adsorbed on top of the mesoporous film. The deposition of the protein was proved by measurements in the contact angle, showing an increment in the hydrophobicity while the protein concentration is higher. By measuring the dehydrogenase activity in MC3T3-E1 cells cultured in dually functionalized mesoporous titatina films with gentamicin and BMP-2 is possible to find an improvement in cell proliferation. For this purpose, the absorption of a yellow-color formazan dye, product of a water-soluble salt (WST-8) reduction by the dehydrogenases, is measured. In summary, this study proves that by means of the surface modification of MTFs with proteins and loading of gentamicin is possible to achieve an antibacterial effect and a cell growth improvement.

Keywords: antibacterial, biocompatibility, bone morphogenetic protein-2, cell proliferation, gentamicin, implants, mesoporous titania films, osteoblasts

Procedia PDF Downloads 163

Authors: Yu-Wen Lin, Chin-Sheng Tang, Wan-Yi Chen

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Various biological specimens, drugs, and chemicals exist in the hospital. The medical staffs and hypersensitive inpatients expose might expose to multiple hazards while they work or stay in the hospital. Therefore, the indoor air quality (IAQ) of the hospital should be paid more attention. Respiratory care wards (RCW) are responsible for caring the patients who cannot spontaneously breathe without the ventilators. The patients in RCW are easy to be infected. Compared to the bacteria concentrations of other hospital units, RCW came with higher values in other studies. This research monitored the IAQ of the RCW and checked the compliances of the indoor air quality standards of Taiwan Indoor Air Quality Act. Meanwhile, the influential factors of IAQ and the impacts of ventilator modules, with humidifier or with filter, were investigated. The IAQ of two five-bed wards and one nurse station of a RCW in a regional hospital were monitored. The monitoring was proceeded for 16 hours or 24 hours during the sampling days with a sampling frequency of 20 minutes per hour. The monitoring was performed for two days in a row and the AIQ of the RCW were measured for eight days in total. The concentrations of carbon dioxide (CO₂), carbon monoxide (CO), particulate matter (PM), nitrogen oxide (NOₓ), total volatile organic compounds (TVOCs), relative humidity (RH) and temperature were measured by direct reading instruments. The bioaerosol samples were taken hourly. The hourly air change rate (ACH) was calculated by measuring the air ventilation volume. Human activities were recorded during the sampling period. The linear mixed model (LMM) was applied to illustrate the impact factors of IAQ. The concentrations of CO, CO₂, PM, bacterial and fungi exceeded the Taiwan IAQ standards. The major factors affecting the concentrations of CO, PM₁ and PM₂.₅ were location and the number of inpatients. The significant factors to alter the CO₂ and TVOC concentrations were location and the numbers of in-and-out staff and inpatients. The number of in-and-out staff and the level of activity affected the PM₁₀ concentrations statistically. The level of activity and the numbers of in-and-out staff and inpatients are the significant factors in changing the bacteria and fungi concentrations. Different models of the patients’ ventilators did not affect the IAQ significantly. The results of LMM can be utilized to predict the pollutant concentrations under various environmental conditions. The results of this study would be a valuable reference for air quality management of RCW.

Keywords: respiratory care ward, indoor air quality, linear mixed model, bioaerosol

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Authors: Kichan Lee, Kwang-Ho Choi, Mi-Hye Hwang, Young Min Son, Bang-Hun Hyun, Byeong Yeal Jung

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Recently, worldwide food safety authorities have been strengthening food hygiene in order to curb foodborne illness outbreaks. The hygiene status of Korean slaughterhouses has been monitored annually by Animal and Plant Quarantine Agency and provincial governments through foodborne pathogens investigation using slaughtered pig and cattle meats. This study presented the prevalence of food-borne pathogens from 2014 to 2016 in Korean slaughterhouses. Sampling, microbiological examinations, and analysis of results were performed in accordance with ‘Processing Standards and Ingredient Specifications for Livestock Products’. In total, swab samples from 337 pig carcasses (100 samples in 2014, 135 samples in 2015, 102 samples in 2016) and 319 cattle carcasses (100 samples in 2014, 119 samples in 2015, 100 samples in 2016) from twenty slaughterhouses were examined for Listeria monocytogenes, Campylobacter jejuni, Campylobacter coli, Salmonella spp., Staphylococcus aureus, Clostridium perfringens, Yersinia enterocolitica, Escherichia coli O157:H7 and non-O157 enterohemorrhagic E. coli (EHEC, serotypes O26, O45, O103, O104, O111, O121, O128 and O145) as foodborne pathogens. The samples were analyzed using cultural and PCR-based methods. Foodborne pathogens were isolated in 78 (23.1%) out of 337 pig samples. In 2014, S. aureus (n=17) was predominant, followed by Y. enterocolitica (n=7), C. perfringens (n=2) and L. monocytogenes (n=2). In 2015, C. coli (n=14) was the most prevalent, followed by L. monocytogenes (n=4), S. aureus (n=3), and C. perfringens (n=2). In 2016, S. aureus (n=16) was the most prevalent, followed by C. coli (n=13), L. monocytogenes (n=2) and C. perfringens (n=1). In case of cattle carcasses, foodborne bacteria were detected in 41 (12.9%) out of 319 samples. In 2014, S. aureus (n=16) was the most prevalent, followed by Y. enterocolitica (n=3), C. perfringens (n=3) and L. monocytogenes (n=2). In 2015, L. monocytogenes was isolated from 4 samples, S. aureus from three, C. perfringens, Y. enterocolitica and Salmonella spp. from one, respectively. In 2016, L. monocytogenes (n=6) was the most prevalent, followed by C. perfringens (n=3) C. jejuni (n=1), respectively. It was found that 10 carcass samples (4 cattle and 6 pigs) were contaminated with two bacterial pathogen tested. Interestingly, foodborne pathogens were more detected from pig carcasses than cattle carcasses. Although S. aureus was predominantly detected in this study, other foodborne pathogens were also isolated in slaughtered meats. Results of this study alerted the risk of foodborne pathogen infection for humans from slaughtered meats. Therefore, the authors insisted that it was important to enhance hygiene level of slaughterhouses according to Hazard Analysis and Critical Control Point.

Keywords: carcass, cattle, foodborne, Korea, pathogen, pig

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Authors: Amr Saeb, Khalid Al-Rubeaan, Mohamed Abouelhoda, Manojkumar Selvaraju, Hamsa Tayeb

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Background: P. mirabilis is a common uropathogenic bacterium that can cause major complications in patients with long-standing indwelling catheters or patients with urinary tract anomalies. In addition, P. mirabilis is a common cause of chronic osteomyelitis in diabetic foot ulcer (DFU) patients. Methodology: P. mirabilis SCDR1 was isolated from a diabetic ulcer patient. We examined P. mirabilis SCDR1 levels of resistance against nano-silver colloids, the commercial nano-silver and silver containing bandages and commonly used antibiotics. We utilized next generation sequencing techniques (NGS), bioinformatics, phylogenetic analysis and pathogenomics in the identification and characterization of the infectious pathogen. Results: P. mirabilis SCDR1 is a multi-drug resistant isolate that also showed high levels of resistance against nano-silver colloids, nano-silver chitosan composite and the commercially available nano-silver and silver bandages. The P. mirabilis-SCDR1 genome size is 3,815,621 bp with G+C content of 38.44%. P. mirabilis-SCDR1 genome contains a total of 3,533 genes, 3,414 coding DNA sequence genes, 11, 10, 18 rRNAs (5S, 16S, and 23S), and 76 tRNAs. Our isolate contains all the required pathogenicity and virulence factors to establish a successful infection. P. mirabilis SCDR1 isolate is a potential virulent pathogen that despite its original isolation site, wound, it can establish kidney infection and its associated complications. P. mirabilis SCDR1 contains several mechanisms for antibiotics and metals resistance including, biofilm formation, swarming mobility, efflux systems, and enzymatic detoxification. Conclusion: P. mirabilis SCDR1 is the spontaneous nano-silver resistant bacterial strain. P. mirabilis SCDR1 strain contains all reported pathogenic and virulence factors characteristic for the species. In addition, it possesses several mechanisms that may lead to the observed nano-silver resistance.

Keywords: Proteus mirabilis, multi-drug resistance, silver nanoparticles, resistance, next generation sequencing techniques, genome analysis, bioinformatics, phylogeny, pathogenomics, diabetic foot ulcer, xenobiotics, multidrug resistance efflux, biofilm formation, swarming mobility, resistome, glutathione S-transferase, copper/silver efflux system, altruism

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Authors: Huseyin Apdik, Aysegul Dogan, Selami Demirci, Ezgi Avsar Apdik, Fikrettin Sahin

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Mesenchymal stem cells (MSCs) are crucial cell types for bone maintenance and growth along with resident bone progenitor cells providing bone tissue integrity during osteogenesis and skeletal growth. Any deficiency in this regulation would result in vital bone diseases. Of those, osteoporosis, characterized by a reduction in bone mass and mineral density, is a critical skeletal disease for especially elderly people. The commonly used drugs for the osteoporosis treatment are bisphosphonates (BPs). The most prominent role of BPs is to prevent bone resorption arisen from high osteoclast activity. However, administrations of bisphosphonates may also cause bisphosphonate-induced osteonecrosis of the jaw (BIONJ). Up to the present, the researchers have proposed several circumstances for BIONJ. However, effects of long-term and/or high dose usage of BPs on stem cell’s proliferation, survival, differentiation or maintenance capacity have not been evaluated yet. The present study will be held to; figure out BPs’ effects on MSCs in vitro in the aspect of cell proliferation and toxicity, migration, angiogenic activity, lineage specific gene and protein expression levels, mesenchymal stem cell properties and potential signaling pathways affected by BP treatment. Firstly, mesenchymal stem cell characteristics of Dental Pulp Stem Cells (DPSCs) and Periodontal Ligament Stem Cells (PDLSCs) were proved using flow cytometry analysis. Cell viability analysis was completed to determine the cytotoxic effects of BPs (Zoledronate (Zol), Alendronate (Ale) and Risedronate (Ris)) on DPSCs and PDLSCs by the 3-(4,5-di-methyl-thiazol-2-yl)-5-(3-carboxymethoxy-phenyl)-2-(4-sulfo-phenyl)-2H-tetrazolium (MTS) assay. Non-toxic concentrations of BPs were determined at 24 h under growth condition, and at 21 days under osteogenic differentiation condition for both cells. The scratch assay was performed to evaluate their migration capacity under the usage of determined of BPs concentrations at 24 h. The results revealed that while the scratch closure is 70% in the control group for DPSCs, it was 57%, 66% and 66% in Zol, Ale and Ris groups, respectively. For PDLSs, while wound closure is 71% in control group, it was 65%, 66% and 66% in Zol, Ale and Ris groups, respectively. As future experiments, tube formation assay and aortic ring assay will be done to determinate angiogenesis abilities of DPSCs and PDLSCs treated with BPs. Expression levels of osteogenic differentiation marker genes involved in bone development will be determined using real time-polymerase change reaction (RT-PCR) assay and expression profiles of important proteins involved in osteogenesis will be evaluated using western blotting assay for osteogenically differentiated MSCs treated with or without BPs. In addition to these, von Kossa staining will be performed to measure calcium mineralization status of MSCs.

Keywords: bisphosphonates, bisphosphonate-induced osteonecrosis of the jaw, mesenchymal stem cells, osteogenesis

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Authors: G. A. Gevorgyan, A. S. Mamyan, L. G. Stepanyan, L. R. Hambaryan

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Bacterio- and phytoplankton growth rates in 'Yerevanyan lich' reservoir and the Hrazdan river in Yerevan city, Armenia were investigated in April and June-August, 2015. Phytoplankton sampling and analysis were performed by the standard methods accepted in hydrobiological studies. The quantitative analysis of aerobic, coliform and E. coli bacteria is done by the 'RIDA COUNT' medium sheets (coated with ready-to-use culture medium). The investigations showed that the insufficient management of household discharges in Yerevan city caused the organic and fecal pollution of the Hrazdan river in this area which in turn resulted in an increase in bacterial count and increased sanitary and pathogenic risks to the environment and human health. During the investigation in April, the representatives of diatom algae prevailed quantitatively in the coastal area of 'Yerevanyan lich' reservoir, nevertheless, a significant change in the phytoplankton community in June occurred: due to green algae bloom in the reservoir, the quantitative parameters of phytoplankton increased significantly. This was probably conditioned by a seasonal increase in the water temperature in the conditions of the sufficient concentration of nutrients. However, a succession in phytoplankton groups during July-August occurred, and a dominant group (according to quantitative parameters) in the phytoplankton community was changed as follows: green algae-diatom algae-blue-green algae. Rapid increase in the quantitative parameters of diatom and blue-green algae in the reservoir may have been conditioned by increased organic matter level resulted from green algae bloom. Algal bloom in 'Yerevanyan lich' reservoir caused changes in phytoplankton community and an increase in bacterioplankton count not only in the reservoir but also in the Hrazdan river sites located in the downstream from the reservoir. Thus, the insufficient management of urban discharges and aquatic ecosystems in Yerevan city led to unfavorable changes in water quality and microbial and phytoplankton communities in “Yerevanyan lich” reservoir and the Hrazdan river which in turn caused increased sanitary and pathogenic risks to the environment and human health.

Keywords: algal bloom, bacterioplankton, phytoplankton, Hrazdan river, Yerevanyan lich reservoir

Procedia PDF Downloads 275

Authors: Temesgen Sidamo, Prakruti S. Rao, Eleni Akllilu, Workineh Shibeshi, Yumi Park, Yong-Soon Cho, Jae-Gook Shin, Scott K. Heysell, Stellah G. Mpagama, Ephrem Engidawork

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The fluoroquinolones (FQs) are used off-label for the treatment of multidrug-resistant tuberculosis (MDR-TB), and for evaluation in shortening the duration of drug-susceptible TB in recently prioritized regimens. Within the class, levofloxacin (LFX) and moxifloxacin (MXF) play a substantial role in ensuring success in treatment outcomes. However, sub-therapeutic plasma concentrations of either LFX or MXF may drive unfavorable treatment outcomes. To the best of our knowledge, the pharmacokinetics of LFX and MXF in Ethiopian patients with MDR-TB have not yet been investigated. Therefore, the aim of this study was to develop a population pharmacokinetic (PopPK) model of levofloxacin (LFX) and moxifloxacin (MXF) and assess the percent probability of target attainment (PTA) as defined by the ratio of the area under the plasma concentration-time curve over 24-h (AUC0-24) and the in vitro minimum inhibitory concentration (MIC) (AUC0-24/MIC) in Ethiopian MDR-TB patients. Steady-state plasma was collected from 39 MDR-TB patients enrolled in the programmatic treatment course and the drug concentrations were determined using optimized liquid chromatography-tandem mass spectrometry. In addition, the in vitro MIC of the patients' pretreatment clinical isolates was determined. PopPK and simulations were run at various doses, and PK parameters were estimated. The effect of covariates on the PK parameters and the PTA for maximum mycobacterial kill and resistance prevention was also investigated. LFX and MXF both fit in a one-compartment model with adjustments. The apparent volume of distribution (V) and clearance (CL) of LFX were influenced by serum creatinine (Scr), whereas the absorption constant (Ka) and V of MXF were influenced by Scr and BMI, respectively. The PTA for LFX maximal mycobacterial kill at the critical MIC of 0.5 mg/L was 29%, 62%, and 95% with the simulated 750 mg, 1000 mg, and 1500 mg doses, respectively, whereas the PTA for resistance prevention at 1500 mg was only 4.8%, with none of the lower doses achieving this target. At the critical MIC of 0.25 mg/L, there was no difference in the PTA (94.4%) for maximum bacterial kill among the simulated doses of MXF (600 mg, 800 mg, and 1000 mg), but the PTA for resistance prevention improved proportionately with dose. Standard LFX and MXF doses may not provide adequate drug exposure. LFX PopPK is more predictable for maximum mycobacterial kill, whereas MXF's resistance prevention target increases with dose. Scr and BMI are likely to be important covariates in dose optimization or therapeutic drug monitoring (TDM) studies in Ethiopian patients.

Keywords: population PK, PTA, moxifloxacin, levofloxacin, MDR-TB patients, ethiopia

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Authors: Ibrahim Siddig Hamid, Ikram Mohamed Eltayeb

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Trachyspermum ammi belongs to the family Apiaceae, is used traditionally for the treatment of gastrointestinal ailments, lack of appetite and bronchial problems as well used as antiseptic, antimicrobial, antipyretic, febrifugal and in the treatment of typhoid fever. Peganum harmala belongs to the family Zygophyllaceae it has been reported to have an antibacterial activity and used to treat depression and recurring fevers. It also used to kill algae, bacteria, intestinal parasites and molds. In Sudan, the combination of two plants are traditionally used for the treatment of bacillary dysentery. Bacillary dysentery is caused by one or more types of Shigella species bacteria mainly Shigella dysenteri and shigella flexneri. Bacillary dysentery is mainly found in hot countries like Sudan with poor hygiene and sanitation. Bacillary dysentery causes sudden onset of high fever and chills, abdominal pain, cramps and bloating, urgency to pass stool, weight loss, and dehydration and if left untreated it can lead to serious complications including delirium, convulsions and coma. A serious infection like this can be fatal within 24 hours. The objective of this study is to investigate the in vitro susceptibility of Sh. flexneri and Sh. dysenteriae to the T. ammi and P. harmala. T. ammi and P. harmala were extracted by 96% ethanol using Soxhlet apparatus. The antimicrobial activity of the extracts was investigated according to the disc diffusion method. The discs were prepared by soaking sterilized filter paper discs in 20 microliter of serially diluted solutions of each plant extract with the concentrations (100, 50, 25, 12.5, 6.25mg/dl) then placing them on Muller Hinton Agar plates that were inoculated with bacterial suspension separately, the plates were incubated for 24 hours at 37c and the minimum inhibitory concentration of the extract which was the least concentration of the extract to inhibit fungal growth was determined. The results showed the high antimicrobial activity of T. ammi extract with an average diameter zone ranging from 18-20 mm and its minimum inhibitory concentration was found to be 25 mg/ml against the two shigella species. P. harmala extract was found to have slight antibacterial effect against the two bacteria. This result justified the Sudanese traditional use of Trachyspermum ammi plant for the treatment of bacillary dysentery.

Keywords: harmala, peganum, shigella, trachyspermum

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Authors: Ghasem Yazdanpanah, Mona Kakavand, Hassan Niknejad

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Objectives: An important issue in tissue engineering (TE) is hemocompatibility. The current engineered vessels are seriously at risk of thrombus formation and stenosis. Amnion (AM) is the innermost layer of fetal membranes that consists of epithelial and mesenchymal sides. It has the advantages of low immunogenicity, anti-inflammatory and anti-bacterial properties as well as good mechanical properties. We recently introduced the amnion as a natural biomaterial for tissue engineering. In this study, we have evaluated hemocompatibility of amnion as potential biomaterial for tissue engineering. Materials and Methods: Amnions were derived from placentas of elective caesarean deliveries which were in the gestational ages 36 to 38 weeks. Extracted amnions were washed by cold PBS to remove blood remnants. Blood samples were obtained from healthy adult volunteers who had not previously taken anti-coagulants. The blood samples were maintained in sterile tubes containing sodium citrate. Plasma or platelet rich plasma (PRP) were collected by blood sample centrifuging at 600 g for 10 min. Hemocompatibility of the AM samples (n=7) were evaluated by measuring of activated partial thromboplastin time (aPTT), prothrombin time (PT), hemolysis, and platelet aggregation tests. P-selectin was also assessed by ELISA. Both epithelial and mesenchymal sides of amnion were evaluated. Glass slide and expanded polytetrafluoroethylene (ePTFE) samples were defined as control. Results: In comparison with glass as control (13.3 ± 0.7 s), prothrombin time was increased significantly while each side of amnion was in contact with plasma (p<0.05). There was no significant difference in PT between epithelial and mesenchymal surfaces (17.4 ± 0.7 s vs. 15.8 ± 0.7 s, respectively). However, aPPT was not significantly changed after incubation of plasma with amnion epithelial and mesenchymal surfaces or glass (28.61 ± 1.39 s, 31.4 ± 2.66 s, glass, 30.76 ± 2.53 s, respectively, p>0.05). Amnion surfaces, ePTFE and glass samples have less hemolysis induction than water considerably (p<0.001), in which no differences were detected. Platelet aggregation measurements showed that platelets were less stimulated by the amnion epithelial and mesenchymal sides, in comparison with ePTFE and glass. In addition, reduction in amount of p-selectin, as platelet activation factor, after incubation of samples with PRP indicated that amnion has less stimulatory effects on platelets than ePTFE and glass. Conclusion: Amnion as a natural biomaterial has the potential to be used in tissue engineering. Our results suggest that amnion has appropriate hemocompatibility to be employed as a vascular substitute.

Keywords: amnion, hemocompatibility, tissue engineering, biomaterial

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Authors: A. Díaz-Roa, P. I. Silva Junior, F. J. Bello

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Sarconesiopsis magellanica (Diptera: Calliphoridae) is a medically important necrophagous fly which is used for establishing the post-mortem interval. Dipterous maggots release diverse proteins and peptides contained in larval excretion and secretion (ES) products playing a key role in digestion. The most important mechanism for combating infection using larval therapy depends on larval ES. These larvae are protected against infection by a diverse spectrum of antimicrobial peptides (AMPs), one already known like lucifensin. Special interest in these peptides has also been aroused regarding understanding their role in wound healing since they degrade necrotic tissue and kill different bacteria during larval therapy. The action of larvae on wounds occurs through 3 mechanisms of action: removal of necrotic tissue, stimulation of granulation tissue, and antibacterial action of larval ES. Some components of the ES include calcium, urea, allantoin ammonium bicarbonate and reducing the viability of Gram positive and Gram negative bacteria. The Lucilia sericata fly larvae have been the most used, however, we need to evaluate new species that could potentially be similar or more effective than fly above. This study was thus aimed at identifying and characterizing S. magellanica AMPs contained in ES products for the first time and compared them with the common fly used L. sericata. These products were obtained from third-instar larvae taken from a previously established colony. For the first analysis, ES fractions were separate by Sep-Pak C18 disposable columns (first step). The material obtained was fractionated by RP-HPLC by using Júpiter C18 semi-preparative column. The products were then lyophilized and their antimicrobial activity was characterized by incubation with different bacterial strains. The first chromatographic analysis of ES from L. sericata gives 6 fractions with antimicrobial activity against Gram-positive bacteria Micrococus luteus, and 3 fractions with activity against Gram-negative bacteria Pseudomonae aeruginosa while the one from S. magellanica gaves 1 fraction against M. luteus and 4 against P. aeruginosa. Maybe one of these fractions could correspond to the peptide already known from L. sericata. These results show the first work for supporting further experiments aimed at validating S. magellanica use in larval therapy. We still need to search if we find some new molecules, by making mass spectrometry and ‘de novo sequencing’. Further studies are necessary to identify and characterize them to better understand their functioning.

Keywords: antimicrobial peptides, larval therapy, Lucilia sericata, Sarconesiopsis magellanica

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Authors: Omaima A. Sharaf, Tarek A. Moussa, Said M. Badr El-Din, H. Moawad

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Background: Polycyclic aromatic hydrocarbons (PAHs) pose great environmental and human health concerns for their widespread occurrence, persistence, and carcinogenic properties. PAHs releases due to anthropogenic activities to the wider environment have led to higher concentrations of these contaminants than would be expected from natural processes alone. This may result in a wide range of environmental problems that can accumulate in agricultural ecosystems, which threatened to become a negative impact on sustainable agricultural development. Thus, this study aimed to evaluate the physico-chemical, and microbial properties of the compost leachate (CL) to assess its role as nutrient and microbial source (biostimulation/bioaugmentation) for developing a cost-effective bioremediation technology for PAHs contaminated sites. Material and Methods: PAHs-degrading bacteria were isolated from CL that was collected from a composting site located in central Scotland, UK. Isolation was carried out by enrichment using phenanthrene (PHR), pyrene (PYR) and benzo(a)pyrene (BaP) as the sole source of carbon and energy. The isolates were characterized using a variety of phenotypic and molecular properties. Six different isolates were identified based on the difference in morphological and biochemical tests. The efficiency of these isolates in PAHs utilization was assessed. Further analysis was performed to define taxonomical status and phylogenic relation between the most potent PAHs-utilizing bacterial strains and other standard strains, using molecular approach by partial 16S rDNA gene sequence analysis. Results indicated that the 16S rDNA sequence analysis confirmed the results of biochemical identification, as both of biochemical and molecular identification of the isolates assigned them to Bacillus licheniformis, Pseudomonas aeruginosa, Alcaligenes faecalis, Serratia marcescens, Enterobacter cloacae and Providenicia which were identified as the prominent PAHs-utilizers isolated from CL. Conclusion: This study indicates that the CL samples contain a diverse population of PAHs-degrading bacteria and the use of CL may have a potential for bioremediation of PAHs contaminated sites.

Keywords: polycyclic aromatic hydrocarbons, physico-chemical analyses, compost leachate, microbial and biochemical analyses, phylogenic relations, 16S rDNA sequence analysis

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Authors: O. Antypenko, I. Vasilieva, S. Kovalenko

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Investigations for new effective and less toxic antimicrobials agents are always up-to-date. The tetrazole derivatives are quite interesting objects as for synthesis as well as for pharmacological screening. Thus, some derivatives of tetrazole demonstrated antimicrobial activity, namely 5-phenyl-tetrazolo[1,5-c]quinazoline was effective one against Staphylococcus aureus and Esherichia faecalis (MIC = 250 mg/L). Besides, investigation of the 9-bromo(chloro)-5-morpholin(piperidine)-4-yl-tetrazolo[1,5-c]quinazoline’s antimicrobial activity against Esherichia coli and Enterococcus faecalis, Pseudomonas aeruginosa and Staphylococcus aureus revealed that sensitivity of Gram-positive bacteria to the compounds was higher than that of Gram-negative bacteria. So, our previously synthesized, 31 derivatives of 1-(2-(1H-tetrazol-5-yl)-R1-phenyl)-3-R2-phenyl(ethyl)ureas were decided to test for their in vitro antibacterial activity against Gram-positive bacteria (Staphylococcus aureus ATCC 25923, Enterobacter aerogenes, Enterococcus faecalis ATCC 29212), Gram-negative bacteria (Pseudomonas aeruginosa ATCC 9027, Escherichia coli ATCC 25922, Klebsiella pneumoniae 68) and antifungal properties against Candida albicans ATCC 885653. Agar-diffusion method was used for determination of the preliminary activity compared to well-known reference antimicrobials. All the compounds were dissolved in DMSO at a concentration of 100 μg/disk, using inhibition zone diameter (IZD, mm) as a measure for the antimicrobial activity. The most active turned to be 3 structures, that inhibited several bacterial strains: 1-ethyl-3-(5-fluoro-2-(1H-tetrazol-5-yl)phenyl)urea (1), 1-(4-bromo-2-(1H-tetrazol-5-yl)-phenyl)-3-(4-(trifluoromethyl)phenyl)urea (2) and 1-(4-chloro-2-(1H-tetrazol-5-yl)phenyl)-3-(3-(trifluoromethyl)phenyl)urea (3). IZM (mm) was 40 (Escherichia coli), 25 (Klebsiella pneumonia) for compound 1; 12 (Pseudomonas aeruginosa), 15 (Staphylococcus aureus), 10 (Enterococcus faecalis) for compound 2; 25 (Staphylococcus aureus), 15 (Enterococcus faecalis) for compound 3. The most sensitive to the activity of the substances were Gram-negative bacteria Pseudomonas aeruginosa. While none of compound effected on Candida albicans. Speaking about, reference drugs: Amikacin (30 µg/disk) showed 27 and Ceftazide (30 µg/disk) 25 against Pseudomonas aeruginosa. That is, unfortunately, higher than studied 1-(2-(1H-tetrazol-5-yl)-R1-phenyl)-3-R2-phenyl(ethyl)ureas. Obtained results will be used for further purposeful optimization of the leading compounds in the more effective antimicrobials because of the ever-mounting problem of microorganism’s resistance.

Keywords: antimicrobial, antifungal, compounds, 1-(2-(1H-tetrazol-5-yl)-R1-phenyl)-3-R2-phenyl(ethyl)ureas

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Authors: Maria L. G. Lourenco, Keylla H. N. P. Pereira, Viviane Y. Hibaru, Fabiana F. Souza, Joao C. P. Ferreira, Simone B. Chiacchio, Luiz H. A. Machado

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Neonatal sepsis is a systemic response of acute infection by bacteria that may lead to high mortality in a litter. This study aims to report a case of sepsis by alpha-hemolytic Streptococcus and Mannheimia haemolytica in neonate dogs. A pregnant, mixed-breed bitch at approximately the 60th day of pregnancy was admitted to the Sao Paulo State University (UNESP) Veterinary Hospital, Botucatu, Sao Paulo, Brazil, and subjected to a c-section due to uterine atony and fetuses no heartbeats on the ultrasound examination. The mother presented leukopenia of 1.6 thousand leukocytes, and there was no other information regarding previous clinical history. Among the offspring, four were stillborn, and five were born alive. On clinical examination, neonates weighed between 312 and 384 grams. Reflexes were present, and the newborn's body temperature was between 89.9 ºF and 96.4 ºF. Neonates also presented clinical signs of neonatal infection: omphalitis, abdomen, and extremities with cyanotic color, hematuria, and diarrhea (meconium). Complementary tests revealed leukopenia. The presence of alpha hemolytic streptococcus and Mannheimia haemolytica was revealed in the bacterial culture. The bacteria were sensitive to cephalosporins and penicillin on the antibiogram. Treatment for sepsis was instituted with the drug ceftriaxone, at a dose of 50 mg per kilogram, administered intravenous (jugular vein). Subsequently administered subcutaneous, every 12 hours, for seven days. Heated fluid therapy was performed, with Ringer lactate, at a dose of 4 ml per 100 grams of weight, intravenous. Heating measures were instituted. Blood plasma was also administered, at a dose of 2 mL per 100 grams of weight, administered subcutaneous, as a source of passive immunity. A maternal milk substitute was instituted, and lactation was discontinued since the mother was unable to nurse due to the infection. The mother was neutered during the c-section and treated with ceftriaxone (50 mg/kg). After seven days, the newborns presented normal clinical signs and no alterations in the hemogram. Early diagnosis and intervention were essential for the survival of these patients.

Keywords: neonatal infection, puppies, bacteria, newborn

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Authors: Recep Kesli, Cengiz Demir, Riza Durmaz, Zekiye Bakkaloglu, Aysegul Bukulmez

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Objective: Acute diarrhea disease in children is a major cause of morbidity worldwide and is a leading cause of mortality, and it is the most common agent responsible for acute gastroenteritis in developing countries. With hospitalized children suffering from acute enteric disease up to 50% of the analyzed specimen were positive for rotavirus. Further molecular surveillance could provide a sound basis for improving the response to epidemic gastroenteritis and could provide data needed for the introduction of vaccination programmes in the country. The aim of this study was to investigate the prevalence of viral etiology of the gastroenteritis in children aged 0-6 years with acute gastroenteritis and to determine predominant genotypes of rotaviruses in the province of Afyonkarahisar, Turkey. Methods: An epidemiological study on rotavirus was carried out during 2016. Fecal samples obtained from the 144 rotavirus positive children with 0-6 years of ages and applied to the Pediatric Diseases Outpatient of ANS Research and Practice Hospital, Afyon Kocatepe University with the complaint of diarrhea. Bacterial agents causing gastroenteritis were excluded by using bacteriological culture methods and finally, no growth observed. Rotavirus antigen was examined by both the immunochromatographic (One Step Rotavirus and Adenovirus Combo Test, China) and ELISA (Premier Rotaclone, USA) methods in stool samples. Rotavirus RNA was detected by using one step real-time reverse transcription-polymerase chain reaction (RT-PCR). G and P genotypes were determined using RT-PCR with consensus primers of VP7 and VP4 genes, followed by semi nested type-specific multiplex PCR. Results: Of the total 144 rotavirus antigen-positive samples with RT-PCR, 4 (2,8%) were rejected, 95 (66%) were examined, and 45 (31,2%) have not been examined for PCR yet. Ninety-one (95,8%) of the 95 examined samples were found to be rotavirus positive with RT-PCR. Rotavirus subgenotyping distributions in G, P and G/P genotype groups were determined as; G1:45%, G2:27%, G3:13%, G9:13%, G4:1% and G12:1% for G genotype, and P[4]:33%, P[8]:66%, P[10]:1% for P genotype, and G1P[8]:%37, G2P[4]:%21, G3P[8]:%10, G4P[8]:%1, G9P[8]:%8, G2P[8]:%3 for G/P genotype . Not common genotype combination were %20 in G/P genotype. Conclusions: This study subscribes to the global agreement of the molecular epidemiology of rotavirus which will be useful in guiding the alternative and application of rotavirus vaccines or effective control and interception. Determining the diversity and rates of rotavirus genotypes will definitely provide guidelines for developing the most suitable vaccine.

Keywords: gastroenteritis, genotyping, rotavirus, RT-PCR

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Authors: Monika Patel, Kazuaki Matsumura

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Synthetic hydrogels, with their unique properties such as porosity, strength, and swelling in aqueous environment, are being used in many fields from food additives to regenerative medicines, from diagnostic and pharmaceuticals to drug delivery systems (DDS). But, hydrogels also have some limitations in terms of homogeneity of drug distribution and quantity of loaded drugs. As an alternate, polymeric micelles are extensively used as DDS. With the ease of self-assembly, and distinct stability they remarkably improve the solubility of hydrophobic drugs. However, presently, combinational therapy is the need of time and so are systems which are capable of releasing more than one drug. And it is one of the major challenges towards DDS to control the release of each drug independently, which simple DDS cannot meet. In this work, we present an amphiphilic polypeptide based micelle hydrogel composite to study the dual drug release for wound healing purposes using Amphotericin B (AmpB) and Curcumin as model drugs. Firstly, two differently charged amphiphilic polypeptide chains were prepared namely, poly L-Lysine-b-poly phenyl alanine (PLL-PPA) and poly Glutamic acid-b-poly phenyl alanine (PGA-PPA) through ring opening polymerization of amino acid N-carboxyanhydride. These polymers readily self-assemble to form micelles with hydrophobic PPA block as core and hydrophilic PLL/PGA as shell with an average diameter of about 280nm. The thus formed micelles were loaded with the model drugs. The PLL-PPA micelle was loaded with curcumin and PGA-PPA was loaded with AmpB by dialysis method. Drug loaded micelles showed a slight increase in the mean diameter and were fairly stable in solution and lyophilized forms. For forming the micelles hydrogel composite, the drug loaded micelles were dissolved and were cross linked using genipin. Genipin uses the free –NH2 groups in the PLL-PPA micelles to form a hydrogel network with free PGA-PPA micelles trapped in between the 3D scaffold formed. Different composites were tested by changing the weight ratios of the both micelles and were seen to alter its resulting surface charge from positive to negative with increase in PGA-PPA ratio. The composites with high surface charge showed a burst release of drug in initial phase, were as the composites with relatively low net charge showed a sustained release. Thus the resultant surface charge of the composite can be tuned to tune its drug release profile. Also, while studying the degree of cross linking among the PLL-PPA particles for effect on dual drug release, it was seen that as the degree of crosslinking increases, an increase in the tendency to burst release the drug (AmpB) is seen in PGA-PPA particle, were as on the contrary the PLL-PPA particles showed a slower release of Curcumin with increasing the cross linking density. Thus, two different pharmacokinetic profile of drugs were seen by changing the cross linking degree. In conclusion, a unique charged amphiphilic polypeptide based micelle hydrogel composite for dual drug delivery. This composite can be finely tuned on the basis of need of drug release profiles by changing simple parameters such as composition, cross linking and pH.

Keywords: amphiphilic polypeptide, dual drug release, micelle hydrogel composite, tunable DDS

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Authors: Zuzana Sedrlova, Eva Slaninova, Ines Fritz, Christina Daffert, Stanislav Obruca

Abstract:

Cyanobacteria are ecologically extremely important phototrophic gram-negative bacteria capable of oxygenic photosynthesis. They synthesize many interesting metabolites such as glycogen, carotenoids, but the most interesting metabolites are polyhydroxyalkanoates (PHA). The main advantage of cyanobacteria is the fact they do not require costly organic substrate and, oppositely, cyanobacteria can fix CO₂. PHA serves primarily as a carbon and energy source and occurs in the form of intracellular granules in bacterial cells. It is possible, PHA helps cyanobacteria to survive stress conditions since increased PHA synthesis was observed during cultivation in stress conditions. PHA is microbial biopolymers that are biodegradable with similar properties as petrochemical synthetic plastics. Production of PHA by heterotrophic bacteria is expensive; for price reduction waste materials as input, materials are used. Positively, cyanobacteria principally do not require organic carbon substrate since they are capable of CO₂ fixation. In this work, we demonstrated that stress conditions lead to the highest obtained yields of PHA in cyanobacterial cultures. Two cyanobacterial cultures from genera Synechocystis were used in this work. Cultivations were performed either in Erlenmayer flask or in tube multicultivator. Multiple stressors were applied on cyanobacterial cultures, and stressors include PHA precursors. PHA precursors are chemical substances and some of them do not occur naturally in the environment. Cultivation with the same PHA precursors in the same concentration led to a 1,6x higher amount of PHA when a multicultivator was used. The highest amount of PHA reached 25 % of PHA in dry cyanobacterial biomass. Both strains are capable of co-polymer synthesis in the presence of their structural precursor. The composition of co-polymer differs in Synechocystis sp. PCC 6803 and Synechocystis salina CCALA 192. Synechocystis sp. PCC 6803 cultivated with γ-butyrolakton accumulated co-polymer of 3-hydroxybutyrate (3HB) and 4-hydroxybutyrate (4HB) the composition of the copolymer was 56 % of 4HB and 44 % of 3HB. The total amount of PHA, as well as yield of biomass, was lower than in control due to the toxic properties of γ-butyrolakton. Funding: This study was partly funded by the project GA19- 19-29651L of the Czech Science Foundation (GACR) and partly funded by the Austrian Science Fund (FWF), a project I 4082-B25. This work was supported by Brno, Ph.D. Talent – Funded by the Brno City Municipality.

Keywords: co-polymer, cyanobacteria, PHA, synechocystis

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Authors: Louise H. Gracioso, Marcela P.G. Baltazar, Ingrid R. Avanzi, Bruno Karolski, Luciana J. Gimenes, Claudio O. Nascimento, Elen A. Perpetuo

Abstract:

Introduction: Higher copper concentrations promote a selection pressure on organisms such as plants, fungi and bacteria, which allows surviving only the resistant organisms to the contaminated site. This selective pressure keeps only the organisms most resistant to a specific condition and subsequently increases their bioremediation potential. Despite the bacteria importance for biosphere maintenance, it is estimated that only a small fraction living microbial species has been described and characterized. Due to the molecular biology development, tools based on analysis 16S ribosomal RNA or another specific gene are making a new scenario for the characterization studies and identification of microorganisms in the environment. News identification of microorganisms methods have also emerged like Biotyper (MALDI / TOF), this method mass spectrometry is subject to the recognition of spectroscopic patterns of conserved and features proteins for different microbial species. In view of this, this study aimed to isolate bacteria resistant to copper present in a Copper Processing Area (Sossego Mine, Canaan, PA) and identifies them in two different methods: Recent (spectrometry mass) and conventional. This work aimed to use them for a future bioremediation of this Mining. Material and Methods: Samples were collected at fifteen different sites of five periods of times. Microorganisms were isolated from mining wastes by culture enrichment technique; this procedure was repeated 4 times. The isolates were inoculated into MJS medium containing different concentrations of chloride copper (1mM, 2.5mM, 5mM, 7.5mM and 10 mM) and incubated in plates for 72 h at 28 ºC. These isolates were subjected to mass spectrometry identification methods (Biotyper – MALDI/TOF) and 16S gene sequencing. Results: A total of 105 strains were isolated in this area, bacterial identification by mass spectrometry method (MALDI/TOF) achieved 74% agreement with the conventional identification method (16S), 31% have been unsuccessful in MALDI-TOF and 2% did not obtain identification sequence the 16S. These results show that Biotyper can be a very useful tool in the identification of bacteria isolated from environmental samples, since it has a better value for money (cheap and simple sample preparation and MALDI plates are reusable). Furthermore, this technique is more rentable because it saves time and has a high performance (the mass spectra are compared to the database and it takes less than 2 minutes per sample).

Keywords: copper mining area, bioremediation, microorganisms, identification, MALDI/TOF, RNA 16S

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Authors: Asrat Agalu Abejew

Abstract:

Background: Antimicrobial resistance (AMR) is a silent tsunami and one of the top global threats to health care and public health. It is one of the common agendas globally and in Ethiopia. Emerging AMR will be a double burden to Ethiopia, which is facing a series of problems from infectious disease morbidity and mortality. In Ethiopia, although there are attempts to document AMR in healthcare institutions, comprehensive and all-inclusive analysis is still lacking. Thus, this study is aimed to determine trends in AMR from 2016-2021. Methods: A retrospective analysis of secondary data recorded in the Amhara Public Health Institute (APHI) from 2016 to 2021 G.C was conducted. Blood, Urine, Stool, Swabs, Discharge, body effusions, and other Microbiological specimens were collected from each study participants, and Bacteria identification and Resistance tests were done using the standard microbiologic procedure. Data was extracted from excel in August 2022, Trends in AMR were analyzed, and the results were described. In addition, the chi-square (X2) test and binary logistic regression were used, and a P. value < 0.05 was used to determine a significant association. Results: During 6 years period, there were 25143 culture and susceptibility tests. Overall, 265 (46.2%) bacteria were resistant to 2-4 antibiotics, 253 (44.2%) to 5-7 antibiotics, and 56 (9.7%) to >=8 antibiotics. The gram-negative bacteria were 166 (43.9%), 155 (41.5%), and 55 (14.6%) resistant to 2-4, 5-7, and ≥8 antibiotics, respectively, whereas 99(50.8%), 96(49.2% and 1 (0.5%) of gram-positive bacteria were resistant to 2-4, 5-7 and ≥8 antibiotics respectively. K. pneumonia 3783 (15.67%) and E. coli 3199 (13.25%) were the most commonly isolated bacteria, and the overall prevalence of AMR was 2605 (59.9%), where K. pneumonia 743 (80.24%), E. cloacae 196 (74.81%), A. baumannii 213 (66.56%) being the most common resistant bacteria for antibiotics tested. Except for a slight decline during 2020 (6469 (25.4%)), the overall trend of AMR is rising from year to year, with a peak in 2019 (8480 (33.7%)) and in 2021 (7508 (29.9%). If left un-intervened, the trend in AMR will increase by 78% of variation from the study period, as explained by the differences in years (R2=0.7799). Ampicillin, Augmentin, ciprofloxacin, cotrimoxazole, tetracycline, and Tobramycin were almost resistant to common bacteria they were tested. Conclusion: AMR is linearly increasing during the last 6 years. If left as it is without appropriate intervention after 15 years (2030 E.C), AMR will increase by 338.7%. A growing number of multi-drug resistant bacteria is an alarm to awake policymakers and those who do have the concern to intervene before it is too late. This calls for a periodic, integrated, and continuous system to determine the prevalence of AMR in commonly used antibiotics.

Keywords: AMR, trend, pattern, MDR

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Authors: Maryam Beiranvand

Abstract:

Multi-drug resistance in various microorganisms has increased globally in many healthcare facilities. Less effective antimicrobial activity of drug therapies for infection control becomes trouble. Since 1980, no new type of antimicrobial drug has been identified, even though combinations of antibiotic drugs have been discovered almost every decade. Between 1981 and 2006, over 70% of novel pharmaceuticals and chemical agents came from natural sources. Microorganisms have yielded almost 22,000 natural compounds. The identification of antimicrobial components from endophytes bacteria could help overcome the threat posed by multi-drug resistant strains. The project aims to analyze and identify antimicrobial peptides isolated from entophytic bacteria and their activity against multidrug-resistant Gram-negative bacteria in South Korea. Endophytic Paenibacillus polymyxa. 4G3 isolated from the plant, Gynura procumbery exhibited considerable antimicrobial activity against Methicillin-resistant Staphylococcus aureus, and Escherichia coli. The Rapid Annotations using Subsystems Technology showed that the total size of the draft genome was 5,739,603bp, containing 5178 genes with 45.8% G+C content. Genome annotation using antiSMASH version 6.0.0 was performed, which predicted the most common types of non-ribosomal peptide synthetase (NRPS) and polyketide synthase (PKS). In this study, diethyl aminoethyl cellulose (DEAEC) resin was used as the first step in purifying for unknown peptides, and then the target protein was identified using hydrophilic and hydrophobic solutions, optimal pH, and step-by-step tests for antimicrobial activity. This crude was subjected to C18 chromatography and elution with 0, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, and 100% methanol, respectively. Only the fraction eluted with 20% -60% methanol demonstrated good antimicrobial activity against MDR E. coli. The concentration of the active fragment was measured by the Brad-ford test, and Protein A280 - Thermo Fisher Scientific at the end by examining the SDS PAGE Resolving Gel, 10% Acrylamide and purity were confirmed. Our study showed that, based on the combined results of the analysis and purification. P polymyxa. 4G3 has a high potential exists for producing novel functions of polymyxin E and bacitracin against bacterial pathogens.

Keywords: endophytic bacteria, antimicrobial activity, antimicrobial peptide, whole genome sequencing analysis, multi -drug resistance gram negative bacteria

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