Search results for: in vitro rumen fermentation

Authors: Prabhavathi Sundaram, Heidi Abrahamse

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In recent years, nanotechnology coupled with photodynamic therapy (PDT) has received considerable attention in terms of improving the effectiveness of drug delivery in cancer therapeutics. The development of functionalized single-walled carbon nanotubes (SWCNTs) has become revolutionary in targeted photosensitizers delivery since it improves the therapeutic index of drugs. The objective of this study was to prepare, characterize and evaluate the potential of functionalized SWCNTs using hyaluronic acid and loading it with photosensitizer and to effectively target colon cancer cells. The single-walled carbon nanotubes were covalently functionalized with hyaluronic acid and the loaded photosensitizer by non-covalent interaction. The photodynamic effect of SWCNTs is detected under laser irradiation in vitro. The hyaluronic acid-functionalized nanocomposites had a good affinity with CD44 receptors, and it avidly binds on to the surface of CACO-2 cells. The cellular uptake of nanocomposites was studied using fluorescence microscopy using lyso tracker. The anticancer activity of nanocomposites was analyzed in CACO-2 cells using different studies such as cell morphology, cell apoptosis, and nuclear morphology. The combined effect of nanocomposites and PDT improved the therapeutic effect of cancer treatment. The study suggested that the nanocomposites and PDT have great potential in the treatment of colon cancer.

Keywords: colon cancer, hyaluronic acid, single walled carbon nanotubes, photosensitizers, photodynamic therapy

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Authors: Vincent Alexander, Rizkita Esyanti

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Leaves of Stevia rebaudiana contain steviol glycoside which mainly comprise of stevioside, a natural sweetener compound that is 100-300 times sweeter than sucrose. Current cultivation method of Stevia rebaudiana in Indonesia has yet to reach its optimum efficiency and productivity to produce stevioside as a safe sugar substitute sweetener for people with diabetes. An alternative method that is not limited by environmental factor is in vitro temporary immersion system (TIS) culture method using recipient for automated immersion (RITA^®) bioreactor. The aim of this research was to evaluate the effect of red LED light induction towards shoot growth and stevioside accumulation in TIS RITA^® bioreactor system, as an endeavour to increase the secondary metabolite synthesis. The result showed that the stevioside accumulation in TIS RITA^® bioreactor system induced with red LED light for one hour during night was higher than that in TIS RITA^® bioreactor system without red LED light induction, i.e. 71.04 ± 5.36 μg/g and 42.92 ± 5.40 μg/g respectively. Biomass growth rate reached as high as 0.072 ± 0.015/day for red LED light induced TIS RITA^® bioreactor system, whereas TIS RITA^® bioreactor system without induction was only 0.046 ± 0.003/day. Productivity of Stevia rebaudiana shoots induced with red LED light was 0.065 g/L medium/day, whilst shoots without any induction was 0.041 g/L medium/day. Sucrose, salt, and inorganic consumption in both bioreactor media increased as biomass increased. It can be concluded that Stevia rebaudiana shoot in TIS RITA^® bioreactor induced with red LED light produces biomass and accumulates higher stevioside concentration, in comparison to bioreactor without any light induction.

Keywords: LED, Stevia rebaudiana, Stevioside, TIS RITA

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Authors: N. L. S. Ahmad, Y. L. Thong, P. Nambiar

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An in vitro study was carried out to evaluate the feasibility of small field of view (FOV) cone beam computed tomography (CBCT) in determining endodontic working length. The objectives were to determine the accuracy of CBCT in measuring the estimated preoperative working lengths (EPWL), endodontic working lengths (EWL) and file lengths. Access cavities were prepared in 27 molars. For each root canal, the baseline electronic working length was determined using an EAL (Raypex 5). The teeth were then divided into overextended, non-modified and underextended groups and the lengths were adjusted accordingly. Imaging and measurements were made using the respective software of the RVG (Kodak RVG 6100) and CBCT units (Kodak 9000 3D). Root apices were then shaved and the apical constrictions viewed under magnification to measure the control working lengths. The paired t-test showed a statistically significant difference between CBCT EPWL and control length but the difference was too small to be clinically significant. From the Bland Altman analysis, the CBCT method had the widest range of 95% limits of agreement, reflecting its greater potential of error. In measuring file lengths, RVG had a bigger window of 95% limits of agreement compared to CBCT. Conclusions: (1) The clinically insignificant underestimation of the preoperative working length using small FOV CBCT showed that it is acceptable for use in the estimation of preoperative working length. (2) Small FOV CBCT may be used in working length determination but it is not as accurate as the currently practiced method of using the EAL. (3) It is also more accurate than RVG in measuring file lengths.

Keywords: accuracy, CBCT, endodontics, measurement

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Authors: Sachin Latiyan, T. S. Sampath Kumar, Mukesh Doble

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Natural polymer based nanofibrous wound dressings have gained increased attention because of their high surface area, bioactivity, biodegradability and resemblance to extracellular matrix. Agarose (a natural polymer) have been used largely for angiogenesis, cartilage formation and wound healing applications. However, electrospinning of agarose is tedious thereby rendering limited studies on fabrication and evaluation of agarose based nanofibrous wound dressings. Thus, present study focuses on the fabrication of agarose (10% w/v)/ polyvinyl alcohol (12% w/v) based multifunctional nanofibrous scaffolds. Zinc citrate (1, 3 and 5% w/w of the polymer) was added as a potential antibacterial agent to combat wound infections. The fabricated scaffolds exhibit ~500% swelling (in phosphate buffer saline) with enhanced mechanical strength which is suitable for most of the wound healing applications. In vitro studies were found to reveal an increased migration and proliferation of L929 mouse fibroblasts with agarose blends w.r.t to the control. The fabricated dressings were found to be effective against both Escherichia coli (Gram-negative) and Staphylococcus aureus (Gram-positive) bacterial strains. Hence, a multifunctional (as provides effective swelling and mechanical support along with antibacterial property), natural product based, eco-friendly scaffold was successfully fabricated to serve as a potential wound dressing material.

Keywords: antibacterial dressings, benign solvent, nanofibrous agarose, biocompatibility, enhanced swelling and mechanical strength, biopolymeric dressings

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Authors: Ehab El-Dabaa, Omnia Ali, Mohamed Abd El-Hady, Ahmed Osman

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Recombinant human interferon alpha 2 (rhIFN-α2) is FDA approved for treatment of some viral and malignant diseases. Approved pegylated rhIFN-α2 drugs have highly improved pharmacokinetics, pharmacodynamics and therapeutic efficiency compared to native protein. In this work, we studied the pegylation of purified properly refolded rhIFN-α2b using linear 20kDa PEG-NHS (polyethylene glycol- N-hydroxysuccinimidyl ester) to prepare pegylated rhIFN-α2b with high stability and activity. The effect of different parameters like rhIFN-α2b final concentration, pH, rhIFN-α2b/PEG molar ratios and reaction time on the efficiency of pegylation (high percentage of monopegylated rhIFN-α2b) have been studied in small scale (100µl) pegylation reaction trials. Study of the percentages of different components of these reactions (mono, di, polypegylated rhIFN-α2b and unpegylated rhIFN-α2b) indicated that 2h is optimum time to complete the reaction. The pegylation efficiency increased at pH 8 (57.9%) by reducing the protein concentration to 1mg/ml and reducing the rhIFN-α2b/PEG ratio to 1:2. Using larger scale pegylation reaction (65% pegylation efficiency), ion exchange chromatography method has been optimized to prepare and purify the monopegylated rhIFN-α2b with high purity (96%). The prepared monopegylated rhIFN-α2b had apparent Mwt of approximately 65 kDa and high in vitro antiviral activity (2.1x10⁷ ± 0.8 x10⁷ IU/mg). Although it retained approximately 8.4 % of the antiviral activity of the unpegylated rhIFN-α2b, its activity is high compared to other pegylated rhIFN-α2 developed by using similar approach or higher molecular weight branched PEG.

Keywords: antiviral activity, rhIFN-α2b, pegylation, pegylation efficiency

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Authors: Nitin Dwivedi, Jigna Shah

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Aim and objective of study: This article indicates a comparison among various generations of dendrimers, a dendrimer is a bioactive material has repetitively branched molecule and used for delivery of various therapeutic active agents. This debut report compares the effect various generations of PPI dendrimers for brain targeting and management of neurodegenerative disorders potential on single platform. This report involves the study of the various mechanism of synthesis ligand anchored various generations PPI dendrimers deliver the drug directly to the CNS, prove their effectiveness in the management of the various neurodegenerative disease. Material and Methods: The Memantine an anti-Alzheimer drug loaded in different generations (3.0G, 4.0G, and 5.0G) of PPI dendrimers which were synthesized were synthesized. The various studies investigate the effect of PPI dendrimers generation on different characteristic parameters i.e. synthesis procedure, drug loading, release behavior, hemolysis profile at different concentration, MRI study for determine the route drug from olfactory transfer, animal model study in vitro, as well as in vivo performance. The outcomes of the investigation indicate drug delivery benefit as well as superior biocompatibility of 4.0G PPI dendrimer over 3.0G and 5.0G dendrimer, respectively. Results and Conclusion: The above study indicate the superiority of in drug delivery system with maximum drug utilization and minimize the drug dose for neurodegenerative disorder over 5.0G PPI dendrimers. So, 4.0G PPI dendrimers are the safe formulations for the symptomatic treatment of the neurodegenerative disorder. The fifth-generation poly(propyleneimine) (PPI) dendrimers, inherent toxicity due to the presence of many peripheral cationic groups is the major issue that limits their applicability.

Keywords: Alzheimer disease, generation, memantine, PPI

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Authors: Yu-Shan Wu, Po-Liang Lai, I-Ming Chu

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Poly(methylmethacrylate)(PMMA) is the most frequently used bone void filler for vertebral augmentation in osteoporotic fracture. PMMA bone cement not only exhibits strong mechanical properties but also can fabricate according to the shape of bone defect. However, the adhesion between the PMMA-based cement and the adjacent bone is usually weak and as PMMA bone cement is inherently bioinert. The combination of bioceramics and polymers as composites may increase cell adhesion and improve biocompatibility. The nano-hydroxyapatite(HAP) not only plays a significant role in maintaining the properties of the natural bone but also offers a favorable environment for osteoconduction, protein adhesion, and osteoblast proliferation. However, defects and cracks can form at the polymer/ceramics interface, resulting in uneven distribution of stress and subsequent inferior mechanical strength. Surface-modified HAP nano-crystals were prepared by chemically grafting poly(ε-caprolactone)(PCL) on surface-modified nano-HAP surface to increase the affinity of polymer/ceramic phases .Thus, incorporation of surface-modified nano-hydroxyapatite (EC-HAP) may not only improve the interfacial adhesion between cement and bone and between nanoparticles and cement, but also increase biocompatibility. In this research, PMMA mixing with 0, 5, 10, 15, 20, 25 and 30 wt% EC-HAP were examined. MC3T3-E1 cells were used for the biological evaluation of the response to the cements in vitro. Morphology was observed using scanning electron microscopy (SEM). Mechanical properties of HAP/PMMA and EC-HAP/PMMA cement were investigated by compression test. Surface wettability of the cements was measured by contact angles.

Keywords: bone cement, biocompatibility, nano-hydroxyapatite, polycaprolactone, PMMA, surface grafting

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Authors: Mazouzi Nourdjihane, K. Boutemak, A. Haddad, Y. Chegreouche

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In the last decades, biodegradable polymers have been considered as one of the most popular options for the delivery of drugs and various conventional doses. The film forming system (FFS) can be used in topical, transdermal, ophthalmic, oral and gastric applications. Recently this system has focused on improving drug delivery, which can promote drug release. In this context, the aim of this study is to create polymeric film-forming systems for the stomach and to evaluate and test their gastroprotective effects, comparing the effects of changes in composition on film characteristics. It uses a plant-derived polyphenol extract extracted from fennel to demonstrate anti-inflammatory activity in the film. The films are made from hydroxypropyl methylcellulose polymer and different types of plastic, glycerol and polyethylene glycol. The ffs properties show that MgO-glycerol-reinforced hydroxypropylmethylcellulose (HPMC-MgO-Gly) is better than that based on MgO-PEG-reinforced hydroxypropylmethylcellulose (HPMC-MgO-PEG). It is durable, has a faster drying time and allows for maximum recovery. Water vapor strength and blowing speed and other additions show another advantage of HPMC-MgO-Gly compared to HPMC-MgO-PEG, indicating good adhesion between the support (top) and film production. In this study, the gastroprotective effect of fennel phenol extract was found, showing that this plant material has a gastroprotective effect on ulcers and that the film can absorb the active substance.

Keywords: film formin system, hydroxypropyl methylcellulose, magnesium oxide, in vivo

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Authors: Yi-Chun Wu, Nai-Yun Chang, Chuan LI

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This study investigates a feasible range of composition for the mixture of gelatin and polyvinyl alcohol to form nanofibers by electrospinning. Gelatin, one of the most available naturally derived hydrogels of amino acids, is a popular choice for food additives, cosmetic ingredients, biomedical implants, or dressing of its non-toxic and biodegradable nature. Nevertheless, synthetic hydrogel polyvinyl alcohol has long been used as a thickening agent for adhesion purposes. Many biomedical devices are also containing polyvinyl-alcohol as a major content, such as eye drops and contact lenses. To discover appropriate compositions of gelatin and polyvinyl-alcohol for electrospun nanofibers, polymer solutions of different volumetric ratios between gelatin and polyvinyl alcohol were prepared for electrospinning. The viscosity, surface tension, pH value, and electrical conductance of polymer solutions were measured. On the nanofibers, the vibrational modes of molecular structures in nanofibers were investigated by Fourier-transform infrared spectroscopy. The morphologies and surface chemical elements of fibers were examined by the scanning electron microscope and the energy-dispersive X-ray spectroscopy. The hydrophilicity of nanofiberswas evaluated by the water contact angles on the surface of the fibers. To further test the biotoxicity of nanofibers, an in-vitro 3T3 fibroblasts culture further tested the biotoxicity of the electrospun nanofibers. Throughstatistical analyses of the experimental data, it is found that the polyvinyl-alcohol rich composition (the volumetric ratio of gelatin/polyvinyl-alcohol < 1) would be a preferable choice for the formation of nanofibers by the current setup of electrospinning. These electrospun nanofibers tend to be hydrophilic with no biotoxicity threat to the 3T3 fibroblasts.

Keywords: gelatin, polyvinyl-alcohol, nanofibers, electrospinning, spin coating

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Authors: Murtada Ahmed Oshi, Jin-Wook Yoo

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Layer-by-layer (LBL) coating has gained popularity for drug delivery of therapeutic drugs. Herein we described a novel approach for enhancing the therapeutic efficiency of the locally administered dexamethasone (Dex) for inflammatory bowel disease (IBD). We utilized a LBL-coating technique on Dex microcrystals (DexMCs) with multiple layers of polyelectrolytes composed of poly (allylamine hydrochloride) (PAH), poly (sodium 4-styrene sulfonate) (PSS) and Eudragit® S100 (ES). The successful deposition of the layers onto DexMCs surfaces were confirmed through zeta potential measurement and confocal laser scanning microscopy. The surface morphology was investigated through scanning electron microscopy. The drug encapsulation efficiency was 95% with a mean particle size of 2 µm and negative surface charge (-40 mV). Moreover, in vitro drug release study showed a minimum release of the drug ( 15%) at an acidic condition during initial first 5 h, followed by sustained-release at an alkaline condition. For in vivo study, LBL-DxMCs were administered orally to ICR mice suffering from dextran sulfate sodium-induced colitis. LBL-DxMCs substantially enhanced anti-IBD activities as compared to DxMCs. Macroscopic, histological and biochemical (tumor necrosis factor-α, interleukin-6 and myeloperoxidase) examinations revealed marked improvements of colitis signs in the mice treated with LBL-DxMCs compared with those treated with DxMCs. Overall, LBL-DxMCs could be a suitable candidate for the treatment of IBD.

Keywords: dexamethasone, inflammatory bowel disease, LBL-coating, polyelectrolytes

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Authors: Saba Atefyekta

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Aim: Control of bacterial bioburden in wounds is an important step for minimizing the risk of wound infection. An antimicrobial hydrogel wound dressing is developed out of soft polymeric hydrogels that contain antimicrobial peptides (AMPs). Such wound dressings can bind and kill all types of bacteria, even the resistance types at the wound site. Methods: AMPs are permanently bonded onto a soft nanostructured polymer via covalent attachment and physical entanglement. This improves stability, rapid antibacterial activity, and, most importantly, prevents the leaching of AMPs. Major Findings: Antimicrobial analysis of antimicrobial hydrogels using in-vitro wound models confirmed >99% killing efficiency against multiple bacterial trains, including MRSA, MDR, E. Coli. Furthermore, the hydrogel retained its antibacterial activity for up to 4 days when exposed to human serum. Tests confirmed no release of AMPs, and it was proven non-toxic to mammalian cells. An in-vivo study on human intact skin showed a significant reduction of bacteria for part of the subject’s skin treated with antibacterial hydrogels. A similar result was detected through a qualitative study in veterinary trials on different types of surgery wounds in cats, dogs, and horses. Conclusions: Antimicrobial hydrogels wound dressings developed by permanent attachment of AMPs can effectively and rapidly kill bacteria in contact. Such antibacterial hydrogel wound dressings are non-toxic and do not release any substances into the wound.

Keywords: antibacterial wound dressing, antimicrobial peptides, post-surgical wounds, infection

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Authors: Cheng-Chih Tsai, Yu-Hsuan Liu, Cheng-Ying Ho, Chun-Chin Huang

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The aim of this study evaluated the in vitro and in vivo antimicrobial activity of selected lactic acid bacteria (LAB) against Uropathogenic Escherichia coli (UPEC) for prevention and amelioration of UTIs. We screened LAB strains with antimicrobial effects on UPEC using a well-diffusion assay, bacterial adherence to the uroepithelium cell line SV-HUC-1 (BCRC 60358), and a coculture inhibition assay. The results showed that the 7 LAB strains (Lactobacillus paracasei, L. salivarius, two Pediococcus pentosaceus strains, two L. plantarum strains, and L. crispatus) and the fermented probiotic products produced by these multi-LAB strains exhibited potent zones of inhibition against UPEC. Moreover, the LAB strains and probiotic products adhered strongly to the uroepithelium SV-HUC-1 cell line. The growth of UPEC strains was also markedly inhibited after co-culture with the LAB strains and probiotic products in human urine. In addition, the enhanced levels of IL-6, IL-8 and lactic acid dehydrogenase were significantly decreased by treatments with the LAB strains and probiotic products in UPEC-induced SV-HUC-1 cells. Furthermore, oral administration of probiotic products reduced the number of viable UPEC in the urine of UPEC-challenged BALB/c mice. Taken together, this study demonstrates that probiotic supplementation may be useful as an adjuvant therapy for the treatment of bacterial-induced urinary tract infections.

Keywords: lactic acid bacterium, SV-HUC-1 uroepithelium, urinary tract infection, uropathogenic Escherichia coli, BALB/c mice

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Authors: S. V. Patil, P. S. Dounde, S. S. Patil

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Emulgels have emerged as one of the most interesting topical delivery system as it has dual release control system that is gel and emulsion. The major objective behind this formulation is delivery of hydrophobic drugs to systemic circulation via skin. In fact presence of a gelling agent in water phase converts a classical emulsion in to emulgel. The emulgel for dermatological use has several favorable properties such as being thixotropic, greaseless, easily spreadable, easily removable, emollient, non-staining, water-soluble, longer shelf life, bio-friendly, transparent and pleasing appearance. Various penetration enhancers can potentiate the effect. So this can be used as better topical drug delivery systems over present conventional systems available in market. Piroxicam is a non-steroidal anti-inflammatory drug that has major problems when administered orally; it is an insoluble drug and has irritant effect on gastro intestinal tract lead to ulceration and bleeding. The aim of this study was to overcoming these problems through preparation of topical emulgel of this drug. Emulgel of Piroxicam was prepared using Carbopol 940 along with mentha oil and clove oil as permeation enhancer. The prepared emulgel were evaluated for their physical appearance, pH determination, viscosity, spreadability, in vitro drug release, ex vivo permeation studies. All the prepared formulations showed acceptable physical properties, homogeneity, consistency, spreadability, viscosity and pH value. The emulgel was found to be stable with respect to physical appearance, pH, rheological properties and drug content at all temperature and conditions for three month.

Keywords: emulgel, piroxicam, menthe oil, clove oil

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Authors: Mihriban Korukluoglu, Goksen Arik, Cagla Erdogan, Selen Kocakoglu

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Kefir is a traditional fermented refreshing beverage which is known for its valuable and beneficial properties for human health. Mainly yeast species, lactic acid bacteria (LAB) strains and fewer acetic acid bacteria strains live together in a natural matrix named “kefir grain”, which is formed from various proteins and polysaccharides. Different microbial species live together in slimy kefir grain and it has been thought that synergetic effect could take place between microorganisms, which belong to different genera and species. In this research, yeast and LAB were isolated from kefir samples obtained from Uludag University Food Engineering Department. The cell morphology of isolates was screened by microscopic examination. Gram reactions of bacteria isolates were determined by Gram staining method, and as well catalase activity was examined. After observing the microscopic/morphological and physical, enzymatic properties of all isolates, they were divided into the groups as LAB and/or yeast according to their physicochemical responses to the applied examinations. As part of this research, the antagonistic/synergistic efficacy of the identified five LAB and five yeast strains to each other were determined individually by disk diffusion method. The antagonistic or synergistic effect is one of the most important properties in a co-culture system that different microorganisms are living together. The synergistic effect should be promoted, whereas the antagonistic effect is prevented to provide effective culture for fermentation of kefir. The aim of this study was to determine microbial interactions between identified yeast and LAB strains, and whether their effect is antagonistic or synergistic. Thus, if there is a strain which inhibits or retards the growth of other strains found in Kefir microflora, this circumstance shows the presence of antagonistic effect in the medium. Such negative influence should be prevented, whereas the microorganisms which have synergistic effect on each other should be promoted by combining them in kefir grain. Standardisation is the most desired property for industrial production. Each microorganism found in the microbial flora of a kefir grain should be identified individually. The members of the microbial community found in the glue-like kefir grain may be redesigned as a starter culture regarding efficacy of each microorganism to another in kefir processing. The main aim of this research was to shed light on more effective production of kefir grain and to contribute a standardisation of kefir processing in the food industry.

Keywords: antagonistic effect, kefir, lactic acid bacteria (LAB), synergistic, yeast

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Authors: Agustine Melviana, Rizkita Esyanti

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Stevia plant contains steviol glycosides which is estimated to be 300 times sweeter than sucrose. However in Indonesia, conventional (in vivo) propagation of Stevia rebaudiana was not effective due to a poor result. Therefore, alternative methods to propagate S. rebaudiana plants is needed, one of it is using in vitro method. Multiplication with a large quantity of stevia biomass in relatively short period can be conducted by using TIS RITA® (Recipient for Automated Temporary Immersion System). The objective of this study was to evaluate the effect of immersion period of the medium on growth and the medium bioconversion into the production of shoot biomass. The study was conducted to determine the effect of different intensity period of medium to enhance biomass of stevia shoots. Shoot culture of S. rebaudiana was grown in full strength MS medium supplemented with 1 ppm Kinetin. RITA® bioreactors were set up with two different immersion periods, 15 min (RITA® 15) and 30 min (RITA® 30), scheduled every 6 hours and incubated for 21 days. The result indicated that immersion period affected the biomass and growth rate (µ). Thirty-minutes immersion showed greater percentage of shoot multiplication (93.44 ± 0.83%), percentage of leaf growth (85.24 ± 5.99%), growth rate (0.042 ± 0.001 g/day), and productivity (0.066 g/L medium/day) compared to that immersed in RITA® 15 min (76.90 ± 4.85%; 79.73 ± 7.76; 0.045 ± 0.004 g/day, and 0.045 g/L medium/day respectively). Enhancement of biomass in RITA® 30 reached 1,702 ± 0,114 gr, whereas in RITA® 15 only 0,953 ± 0,093 gr. Additionally, the pattern of sucrose, mineral, and inorganic compounds consumption followed the growth of plant biomass for both systems. In conclusion, the bioconversion efficiency from medium to biomass in RITA® 30 is better than RITA® 15.

Keywords: intensity period, shoot culture, Stevia rebaudiana, TIS RITA®

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Authors: Mohamed Majrashi, Patricia Connolly, Terry Gourlay

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Cardiopulmonary bypass is known to cause inflammatory response during open heart surgery. It includes the initiation of different cascades such as coagulation, complement system and cytokines. Although the immune system is body’s key defense mechanism against external assault, when overexpressed, it can be injurious to the patient, particularly in a cohort of patients in which there is a heightened and uncontrolled response. The inflammatory response develops in these patients to an exaggerated level resulting in an autoimmune injury and may lead to poor postoperative outcomes (systemic inflammatory response syndrome and multi-organs failure). Previous studies by this group have suggested a correlation between the level of IL6 measured in patient’s blood before surgery and after polymeric activation and the observed inflammatory response during surgery. Based upon these findings, the present work is aimed at using this response to develop a test which can be used prior to the open heart surgery to identify the high-risk patients before their operation. The work will be accomplished via three main clinical phases including some pilot in-vitro studies, device development and clinical investigation. Current findings from studies using animal blood, employing DEHP and DEHP plasticized PVC materials as the activator, support the earlier results in patient samples. Having established this relationship, ongoing work will focus on developing an activated lateral flow strip technology as a screening device for heightened inflammatory propensity.

Keywords: cardiopulmonary bypass, cytokines, inflammatory response, overexpression

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Authors: Senthil Rajan Dharmalingam, Shamala Nadaraju, Srinivasan Ramamurthy

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Doxorubicin is the most widely used anticancer drugs in chemotherapy treatment. However, problems related to the development of multidrug resistance (MDR) and acute cardiotoxicity have led researchers to investigate alternative forms of administering doxorubicin for cancer therapy. Several methods have been attempted to overcome MDR, including the co-administration of a chemosensitizer inhibiting the efflux caused by ATP binding cassette transporters with anticancer drugs, and the bypass of the efflux mechanism. Co encapsulation of doxorubicin (Dox) and cyclosporine A (CSA) into poly (DL-lactide-co-glycolide) nanoparticles was emulsification-solvent evaporation method using polyvinyl alcohol as emulsion stabilizers. The Dox-CSA loaded nanoparticles were evaluated for particle size, zeta potential and PDI by light scattering analysis and thermal characterizations by differential scanning calorimetry (DSC). Loading efficiency (LE %) and in-vitro dissolution samples were evaluated by developed and validated HPLC method. The optimum particle size obtained is 298.6.8±39.4 nm and polydispersity index (PDI) is 0.098±0.092. Zeta potential is found to be -29.9±4.23. Optimum pH to increase Dox LE% was found 7.1 which gave 42.5% and 58.9% increase of LE% for pH 6.6 and pH 8.6 compared respectively. LE% achieved for Dox is 0.07±0.01 % and CSA is 0.09±0.03%. Increased volume of PVA and weight of PLGA shows increase in size of nanoparticles. DSC thermograms showed shift in the melting peak for the nanoparticles compared to Dox and CSA indicating encapsulation of drugs. In conclusion, these preliminary studies showed the feasibility of PLGA nanoparticles to entrap Dox and CSA and require future in-vivo studies to be performed to establish its potential.

Keywords: doxorubicin, cyclosporine, PLGA, nanoparticles

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Authors: Hsin-Yueh Chang, Pin-Chen Liao, Jo-Shu Chang, Chun-Yen Chen

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Biofuel has drawn much attention as a potential substitute to fossil fuels. However, biodiesel from waste oil, oil crops or other oil sources can only satisfy partial existing demands for transportation. Due to the feature of being clean, green and viable for mass production, using microalgae as a feedstock for biodiesel is regarded as a possible solution for a low-carbon and sustainable society. In particular, Thraustochytrium sp. BM2, an indigenous heterotrophic microalga, possesses the potential for metabolizing glycerol to produce lipids. Hence, it is being considered as a promising microalgae-based oil source for biodiesel production and other applications. This study was to optimize the culture pH, scale up, assess the feasibility of producing microalgal lipid from crude glycerol and apply operation strategies following optimal results from shake flask system in a 5L stirred-tank fermenter for further enhancing lipid productivities. Cultivation of Thraustochytrium sp. BM2 without pH control resulted in the highest lipid production of 3944 mg/L and biomass production of 4.85 g/L. Next, when initial glycerol and corn steep liquor (CSL) concentration increased five times (50 g and 62.5 g, respectively), the overall lipid productivity could reach 124 mg/L/h. However, when using crude glycerol as a sole carbon source, direct addition of crude glycerol could inhibit culture growth. Therefore, acid and metal salt pretreatment methods were utilized to purify the crude glycerol. Crude glycerol pretreated with acid and CaCl₂ had the greatest overall lipid productivity 131 mg/L/h when used as a carbon source and proved to be a better substitute for pure glycerol as carbon source in Thraustochytrium sp. BM2 cultivation medium. Engineering operation strategies such as fed-batch and semi-batch operation were applied in the cultivation of Thraustochytrium sp. BM2 for the improvement of lipid production. In cultivation of fed-batch operation strategy, harvested biomass 132.60 g and lipid 69.15 g were obtained. Also, lipid yield 0.20 g/g glycerol was same as in batch cultivation, although with poor overall lipid productivity 107 mg/L/h. In cultivation of semi-batch operation strategy, overall lipid productivity could reach 158 mg/L/h due to the shorter cultivation time. Harvested biomass and lipid achieved 232.62 g and 126.61 g respectively. Lipid yield was improved from 0.20 to 0.24 g/g glycerol. Besides, product costs of three kinds of operation strategies were also calculated. The lowest product cost 12.42 $NTD/g lipid was obtained while employing semi-batch operation strategy and reduced 33% in comparison with batch operation strategy.

Keywords: heterotrophic microalga Thrasutochytrium sp. BM2, microalgal lipid, crude glycerol, fermentation strategy, biodiesel

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Authors: Felicia Segeth, Florian G. Klein, Lea Berger, Andreas Kolk, Per S. Holm

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The entry of oncolytic viruses (OVs) into clinical application opens groundbreaking changes in current and future treatment regimens. However, despite their potent anti-cancer activity in vitro, clinical studies revealed limitations of OVs as monotherapy. The same applies to CDK 4/6 inhibitors (CDK4/6i) targeting cell cycle as well as bromodomain and extra-terminal domain inhibitors (BETi) targeting gene expression. In this study, the anti-tumoral effect of XVir-N-31, an YB-1 dependent oncolytic adenovirus, was evaluated in combination with Ribociclib, a CDK4/6i, and JQ1, a BETi. The head and neck squamous cell carcinoma (HNSCC) cell lines Fadu, SAS, and Cal-33 were used. DNA replication and gene expression of XVir-N-31 was measured by RT-qPCR, protein expression by western blotting, and cell lysis by SRB assays. Treatment with CDK4/6i and BETi increased viral gene expression, viral DNA replication, and viral particle formation. The data show that the combination of oncolytic adenovirus XVir-N-31 with CDK4/6i & BETi acts highly synergistic in cancer cell lysis. Furthermore, additional molecular analyses on this subject demonstrate that the positive transcription elongation factor P-TEFb plays a decisive role in this regard, indicating an influence of the combinational therapy on gene transcription control. The combination of CDK4/6i & BETi and XVir-N-31 is an attractive strategy to achieve substantial cancer cell killing and is highly suitable for clinical testing.

Keywords: adenovirus, BET, CDK4/6, HNSCC, P-TEFb, YB-1

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Authors: Sirikwan Ponprateep, Anchalee Tassanakajon, Vichien Rimphanitchayakit

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A Kazal-type serine proteinase inhibitor, SPIPm2, was abundantly expressed in the hemocytes and secreted into shrimp plasma has anti-viral property against white spot syndrome virus (WSSV). To discover the molecular mechanism of antiviral activity, the binding assay showed that SPIPm2 bind to the components of viral particle and shrimp hemocyte. From our previous report, viral target protein of SPIPm2 was identified, namely WSV477 using yeast two-hybrid screening. WSV477 is an early gene product of WSSV and involved in viral propagation. In this study, the co-immunoprecipitation technique and Tandem Mass Spectrometry (LC-MS/MS) was used to identify the target protein of SPIPm2 from shrimp hemocyte. The target protein of SPIPm2 was cyclophilin A. In vertebrate, cyclophilin A or peptidylprolyl isomerase A was reported to be the immune suppressor interacted with cyclosporin A involved in immune defense response. The recombinant cyclophilin A from Penaeus monodon (rPmCypA) was produced in E.coli system and purified using Ni-NTA column to confirm the protein-protein interaction. In vitro pull-down assay showed the interaction between rSPIPm2 and rPmCypA. To study the biological function of these proteins, the expression analysis of immune gene in shrimp defense pathways will be investigated after rPmCypA administration.

Keywords: cyclophilin A, protein-protein interaction, Kazal-type serine proteinase inhibitor, Penaeus monodon

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Authors: Uche C. Okafor, Nwanneka M. Okpokwu, Felix Nwafor, Carl E. A. Okezie

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Securidaca longepedunculata Fresen (Violet tree) belongs to the family Polygalaceae characterised by papillionaceous purplish flowers. This medicinally valued plant disappears at an alarming rate due to intensified anthropopressure particularly the unregulated manner of subterranean plant parts' collection from natural stands. Some indiscriminately harvested plants bear seeds containing both mature and immature zygotic embryos that are often discarded. Here, such seeds are collected for this experiment. Seeds were collected, washed, de-coated, and dipped in 70 % (v/v) ethanol for 30 s followed by rising in 5 % solution sodium hypochlorite, containing two drops of tween 20, for another 25 min. Mature zygotic embryos (MZEs) were excised from seeds and cultured in two basal media (MS and B5), three carbon sources (sucrose, glucose and fructose) at five concentrations (0-40 g/L) while immature zygotic embryos (iMZEs) were composed on similar basal media and carbon source supplemented with 0-2 mg/L Benzylaminopurine (BAP) and 0-2 mg/L Indole acetic acid (IAA). MZEs cultured on MS + 30g/L sucrose differed significantly from other treatments at p≤0.05 with maximum percent sprouting (85.24± 5.67 %) and shoot length (7.53±0.67 cm). MZEs culture had the maximum percent sprouting (85.24± 5.67 %) and shoot length (7.53±0.67 cm) in medium containing MS+ 30g L-1 sucrose. iMZEs on the other hand had maximum growth on MS + 40g/L sucrose supplemented with 1.5 mg/L IAA+ 1.0 mg/L BAP. This study is a geared towards creating an alternative path for the maximum production of plants in vitro, thereby, preventing the plants from disappearing.

Keywords: Gamborg's medium, Murashige and Skoog medium, Securidaca longepedunculata, zygotic embryos

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Authors: Monthon Tangjitmungman

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Numerous diseases have been linked to oxidative stress, in which a disproportion of free radicals in the body leads to tissue or cell damage. Polyphenols are the most abundant antioxidants found in plants, and they are highly effective at scavenging oxidative free radicals. Due to the presence of phenolic compounds in Caesalpinia sappan has been discovered to have antioxidant activity. It has several health benefits, the most important of which is preventing cardiovascular and cancer diseases. This study aimed to determine the phenolic content and antioxidant activity of C. sappan extract using a variety of antioxidant assays. The extract of C. sappan was made using a mixture of solvents (ethyl alcohol: water in ratio 8:2). The total phenolic content of C. sappan extract was determined and expressed as gallic acid equivalents using the Folin-Cioucalteu method (GAE). The antioxidant activity of C. sappan extract was assessed using the 2, 2-diphenyl-1-picrylhydrazyl (DPPH) free radical scavenging assay and the ABTS radical scavenging capacity assay. An association was found between antioxidant activity and total phenol content. The antioxidant activity of C. sappan extract was also determined by DPPH and ABTS assays. The IC50 values for C. sappan extract from DPPH and ABTS assays were 54.48 μg/mL ± 0.545 and 25.46 μg/mL ± 0.790, respectively, in the DPPH assay. In the DPPH assay, vitamin C was used as a positive control, whereas Trolox was used as a positive control in the ABTS assay. In conclusion, C. sappan extract contains a high level of total phenolics and exhibits significant antioxidant activity. Nevertheless, more research should be done on the antioxidant activity, such as SOD and ROS scavenging assays and in vivo experiments, to determine whether the compound has antioxidant activity.

Keywords: ABTS assay, antioxidant activity, Caesalpinia sappan, DPPH assays, total phenolic content

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Authors: Tuba Kizilirmak

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Distinct properties of porous metamaterials have been largely processed for biomedicine requiring a three-dimensional (3D) porous structure engaged with fine mechanical features, biodegradation ability, and biocompatibility. Applications of metamaterials are (i) porous orthopedic and dental implants; (ii) in vitro cell culture of metamaterials and bone regeneration of metamaterials in vivo; (iii) macro-, micro, and nano-level porous metamaterials for sensors, diagnosis, and drug delivery. There are some specific properties to design metamaterials for tissue engineering. These are surface to volume ratio, pore size, and interconnection degrees are selected to control cell behavior and bone ingrowth. In this study, additive manufacturing technique selective laser melting will be used to print the scaffolds. Selective Laser Melting prints the 3D components according to designed 3D CAD models and manufactured materials, adding layers progressively by layer. This study aims to design metamaterials with Ti6Al4V material, which gives benefit in respect of mechanical and biological properties. Ti6Al4V scaffolds will support cell attachment by conferring a suitable area for cell adhesion. This study will control the osteoblast cell attachment on Ti6Al4V scaffolds after the determination of optimum stiffness and other mechanical properties which are close to mechanical properties of bone. Before we produce the samples, we will use a modeling technique to simulate the mechanical behavior of samples. These samples include different lattice models with varying amounts of porosity and density.

Keywords: additive manufacturing, titanium lattices, metamaterials, porous metals

Procedia PDF Downloads 186

Authors: Jose A. Aznar-Moreno, Xi Jiang, Stefan Burén, Luis M. Rubio

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Integration of prokaryotic nitrogen fixation (nif) genes into the plastid genome for expression of functional nitrogenase components could render plants capable of assimilating atmospheric N2 making their crops less dependent of nitrogen fertilizers. The nitrogenase Fe protein component (NifH) has been used as proxy for expression and targeting of Nif proteins within plant and yeast cells. Here we use tobacco plants with the Azotobacter vinelandii nifH and nifM genes integrated into the plastid genome. NifH and its maturase NifM were constitutively produced in leaves, but not roots, during light and dark periods. Nif protein expression in transplastomic plants was stable throughout development. Chloroplast NifH was soluble, but it only showed in vitro activity when isolated from leaves collected at the end of the dark period. Exposing the plant extracts to elevated temperatures precipitated NifM and apo-NifH protein devoid of [Fe4S4] clusters, dramatically increasing the specific activity of remaining NifH protein. Our data indicate that the chloroplast endogenous [Fe-S] cluster biosynthesis was insufficient for complete NifH maturation, albeit a negative effect on NifH maturation due to excess NifM in the chloroplast cannot be excluded. NifH and NifM constitutive expression in transplastomic plants did not affect any of the following traits: seed size, germination time, germination ratio, seedling growth, emergence of the cotyledon and first leaves, chlorophyll content and plant height throughout development.

Keywords: NifH, chloroplast, nitrogen fixation, crop improvement, transplastomic plants, fertilizer, biotechnology

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Authors: O. M. M. Sabry, Abeer M. El Sayed

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The essential oils from the aerial parts and flowers of Haplophyllum tuberculatum (Forsskal) Adr. Juss (Rutaceae) growing in Libya were obtained separately by hydro-distillation using a Clevenger-type apparatus. The essential oils yield were (0.4, 1.5w/w%) respectively based on the dry weight of the plant. The oils were analyzed by GC-MS. Twenty four constituents, amounting to 96.6%, were identified in the oil of the aerial parts. The predominant compounds were among the non oxygenated terpenoids (82.4%) as monoterpene hydrocarbons, represented by sabinen (26.4 %), δ-terpinen (26 %), β-phellandrene (10.4%) and 3-carene (3.86%). Zingiberine (0.4%) and β-sesquiphellandrene (0.12%) were the major sesquiterpene hydrocarbons identified. Oxygenated monoterpenes were represented by eucalyptol (5.5%) and piperitone (5.55%). Twenty six constituents, equivalent to 99.5%, were identified in the oil of the flowers. The dominance of monoterpene hydrocarbons in the flowers oil can be attributed to the high percentage of γ-terpinen (38.44%), β-phellandrene (10.0%), α- phellandrene (2.33%), 3,4-dimethyl-1,5-cyclooctadiene (6.67%), β-myrecene (6.04%), 3-carene (5.43%) and α-pinene (1.3%).While the oxygenated monoterpenes can be contributed to the trans-piperitol (4.67%) and piperitone (2.07%). Sesquiterpene hydrocarbons were not identified in the oil of the flower of H. tuberculatum. Variation in constitution between oils of Libyan H. tuberculatum and that obtained from other countries can be due to both environmental and genetic factors. The essential oils have demonstrated variable antimicrobial activities against certain micro-organisms. Also have revealed marked in vitro cytotoxicity against lung (H1299), liver (HEPG2) carcinoma cell line and variably effective as anti-inflammatory and antioxidant.

Keywords: Halpophyllum tuberculatum, rutaceae, essential oil, antimicrobial, anti-inflammatory, antitumor, antioxidant, Libya

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Authors: Rahul Madathiparambil Visalakshan, Alex Cavallaro, John Hayball, Krasimir Vasilev

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The role of biomaterial surface nanotopograhy on fibrinogen adsorption and unfolding, and the subsequent immune response were studied. Inconsistent topography and varying chemical functionalities along with a lack of reproducibility pose a challenge in determining the specific effects of nanotopography or chemistry on proteins and cells. It is important to have a well-defined nanotopography with a homogeneous chemistry to study the real effect of nanotopography on biological systems. Therefore, we developed a technique that can produce well-defined and highly reproducible topography to identify the role of specific roughness, size, height and density with the presence of homogeneous chemical functionality. Using plasma polymerisation of oxazoline monomers and immobilized gold nanoparticles we created surfaces with an equal number density of nanoparticles of different sizes. This surface was used to study the role of surface nanotopography and the interplay of surface chemistry on proteins and immune cells. The effect of nanotopography on fibrinogen adsorption was investigated using Quartz Cristal Microbalance with Dissipation and micro BCA. The mass of fibrinogen adsorbed on the surface increased with increasing size of nano-topography. Protein structural changes up on adsorption to the nano rough surface was studied using circular dichroism spectroscopy. Fibrinogen unfolding varied depending on the specific nanotopography of the surfaces. It was revealed that the in vitro immune response to the nanotopography surfaces changed due to this protein unfolding.

Keywords: biomaterial inflammation, protein and cell responses, protein unfolding, surface nanotopography

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Authors: Kriyapa lairungruang, Arunporn Itharat

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Limonia acidissima L. (LA) (Common name: wood apple, Thai name: ma-khwit) is a medicinal plant which has long been used in Thai traditional medicine. Its bark is used for treatment of diarrhea, abscess, wound healing and inflammation and it is also used in oral cancer. Thus, this research aimed to investigate antioxidant and cytotoxic activities of the LA bark extracts produced by various extraction methods. Different extraction procedures were used to extract LA bark for biological activity testing: boiling in water, maceration with 95% ethanol, maceration with 50% ethanol and water boiling of each the 95% and the 50% ethanolic residues. All extracts were tested for antioxidant activity using DPPH radical scavenging assay, cytotoxic activity against human laryngeal epidermoid carcinoma (HEp-2) cells and human oral epidermoid carcinoma (KB) cells using sulforhodamine B (SRB) assay. The results found that the 95% ethanolic extract of LA bark showed the highest antioxidant activity with EC50 values of 29.76±1.88 µg/ml. For cytotoxic activity, the 50% ethanolic extract showed the best cytotoxic activity against HEp-2 and KB cells with IC50 values of 9.55±1.68 and 18.90±0.86 µg/ml, respectively. This study demonstrated that the 95% ethanolic extract of LA bark showed moderate antioxidant activity and the 50% ethanolic extract provided potent cytotoxic activity against HEp-2 and KB cells. These results confirm the traditional use of LA for the treatment of oral cancer and laryngeal cancer, and also support its ongoing use.

Keywords: antioxidant activity, cytotoxic activity, Laryngeal epidermoid carcinoma, Limonia acidissima L., oral epidermoid carcinoma

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Authors: Siti Helmyati, Euis Nurdiyawati, Joko Susilo, Endri Yuliati, Siti Fadhilatun Nashriyah, Kurnia Widyastuti

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Background: Iron fortification is one of the most effective and sustainable strategies to overcome anemia. It contradictively, has negative effect on gut microbiota balance. Pathogenic bacteria required iron for their growth. The iron source have greatly affect iron absorption in the intestine. Probiotic can inhibit the growth of pathogen. Lactobacillus plantarum Dad 13, Indonesian local isolate provides many benefits for health while fructo-oligosaccharides (FOS) provides selective substrates for probiotics’ growth. Objective: To determine the effect of iron fortification (NaFeEDTA and FeSO4) on antibacterial activity of synbiotic fermented milk. Methods: The antibacterial activity test was performed using the disc diffusion method. Paper discs were soaked in three kinds of synbiotic fermented milk, which are: 1) fortified with NaFeEDTA, 2) FeSO4 and 3) control. Escherichia coli was inoculated on nutrient agar medium. The ability of inhibition was shown by the formation of clear zone around the paper disc and measured in diameter (mm). Results: Synbiotic fermented milk fortified with iron (either NaFeEDTA or FeSO4) had antibacterial activity against Escherichia coli with diameter of clear zone were 6.53 mm and 12.3 mm, respectively (p<0.05). Compared to control (10.73 mm), synbiotic fermented milk fortified with FeSO4 had similar antibacterial activity (p>0.05). Conclusions: In vitro, synbiotic fermented milk fortified with NaFeEDTA and FeSO4 had different antibacterial activity against Escherichia coli. Iron fortification compound affected the antibacterial activity of synbiotic fermented milk.

Keywords: lactobacillus plantarum Dad 13, FOS, NaFeEDTA, FeSO4, antibacterial activity

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Authors: Suthida Panwong, Jeeraporn Pekkoh, Yingmanee Tragoolpua, Aussara Panya

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Dengue virus (DENV) infection is a major public health problem in many countries, especially in tropical and subtropical countries. DENV infection causes dengue fever that can progress to serious conditions of dengue hemorrhagic fever (DHF) or dengue shock syndrome (DSS), relevant to a high risk of mortality. However, there are no effective treatments available against the manifestation and fatalities. Currently, natural extracts have been widely used for the treatment of infectious diseases due to their safety, non-accumulation in the body, or lower side effects. Chlorella spp. is a microalgae with anti-viral activity, but there is not much report to support its ability to inhibit DENV infection. Thus, this study aimed to investigate the inhibitory effect of ethanolic extract from Chlorella sp. AARL G049, which was explored in Thailand on inhibition of DENV-2 infection. The inhibitory effect on viral infection was assessed using a foci-forming assay (FFA), which revealed that a concentration of 125 µg/mL could inhibit viral infection in Vero cells by 75.45±8.06% when treated at the same time as DENV-2 infection. Moreover, the extract at an equal concentration effectively reduced viral protein synthesis by 90.51±5.48% when assessed in human cell lines using enzyme-linked immunosorbent assay (ELISA). Concordantly, the number of infected cells after treatment was reduced as measured by immunofluorescent assay (IFA). Therefore, the finding of this study supports the potential use of Chlorella sp. extract to suppress DENV infection.

Keywords: viral infection, flavivirus, treatment, natural extract

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Authors: Kieren Luellman, Makenzi Rockwell, Eduardo Callegari, Nichole Haag, Chun Wu

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Multiple sclerosis (MS) is an autoimmune disorder that damages the myelin sheath of neurons in the central nervous system. The presence of Acinetobacter bacteria and anti-Acinetobacter antibodies in MS patients has led to the hypothesis that the bacteria may contribute to MS pathogenesis. In this study, the protein sequences of Acinetobacter baumannii were compared to five peptides from three mammalian myelin proteins, i.e., Proteolipid Protein (PLP): PLP 139-151, PLP 178-191, Myelin Basic Protein (MBP): MBP 84-104 and Myelin Oligodendrocyte Glycoprotein (MOG): MOG 35-55 and MOG 92-106 respectively, known to induce experimental autoimmune encephalomyelitis (EAE), a condition similar to MS. We found 11 hits (i.e., with five or more amino acid sequence similarity) in Acinetobacter baumannii, which are identical or similar to PLP139-151, 32 hits to PLP178-191, 35 to MBP 84-104, 41 hits to MOG 35-55 and 26 hits to MOG92-106. In addition, Western blotting was used to assess possible interaction between the bacterial proteins and human anti-MBP, anti-MOG, and anti-PLP antibodies produced in rabbits, corresponding to MBP 84-104, MOG 35-55, and PLP 139-151, respectively. We found that both human Polyclonal anti-MOG antibody and anti-PLP antibody recognized a protein or more proteins of the same molecular mass of around 25 kDa. in Acinetobacter baumannii. The results suggested that this/these protein(s) might potentially serve as antigen(s) to induce anti-MOG antibody and anti-PLP antibody production in mammalian B cells. The proteomic study identified 433 hits, among which the sequence of Acinetobacter baumannii protein 491 subunit A matches a previously published enzyme Acinetobacter 3-Oxoadipate CoA-Transferase, in which a fragment of its peptide was observed to recognize MS patient serum via ELISA method. Our findings might pave the road to understanding one of the pathogenesis mechanisms of MS.

Keywords: multiple sclerosis, pathogenesis, Acinetobacter baumannii, antibody recognition

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