Search results for: multi–drug resistant pathogenic bacteria

Authors: Patcharakamon Nooeaid, Piyachat Chuysrinuan

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Electrospun natural polymers-based nanofibers are one of the most interesting materials used in tissue engineering and drug delivery applications. Bead-on-string nanofibers have gained considerable interest for sustained drug release. Vancomycin was used as the model drug and sodium alginate (SA)/soy protein isolated (SPI) as the polymer blend to fabricate the bead-on-string nanofibers by aqueous-based electrospinning. The bead-on-string SA/SPI nanofibers were successfully fabricated by the addition of poly(ethylene oxide) (PEO) as a co-blending polymer. SA-PEO with mass ratio of 70/30 showed the best spinnability with continuous nanofibers without the occurrence of beads. Bead structure formed with the addition of SPI and bead number increased with increasing SPI content. The electrospinning of 80/20 SA-PEO/SPI was obtained as a great promising bead-on-string nanofibers for drug loading, while the solution of 50/50 was not able to obtain continuous fibers. In vitro release tests showed that a more sustainable release profile up to 14 days with less initial burst release on day 1 could be obtained from the bead-on-string fibers than from smooth fibers with uniform diameter. In addition, vancomycin-loaded beaded fibers inhibited the growth of Staphylococcus aureus (S. aureus) bacteria. Therefore, the SA-PEO/SPI nanofibers showed the potential to be used as biomaterials for tissue engineering and drug delivery.

Keywords: bead-on-string fibers, electrospinning, drug delivery, tissue engineering

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Authors: Kalaumari Mayoral-Peña, Ana I. Montejano-Montelongo, Josué Reyes-Muñoz, Gonzalo A. Ortiz-Mancilla, Mayrin Rodríguez-Cruz, Víctor Hernández-Villalobos, Jesús A. Guzmán-López, Santiago García-Jacobo, Iván Licona-Vázquez, Grisel Fierros-Romero, Rosario Flores-Vallejo

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The group B-lactamic antibiotics include some of the most frequently used small drug molecules against bacterial infections. Nevertheless, an alarming decrease in their efficacy has been reported due to the emergence of antibiotic-resistant bacteria. Infections caused by bacteria expressing extended Spectrum B-lactamases (ESBLs) are difficult to treat and account for higher morbidity and mortality rates, delayed recovery, and high economic burden. According to the Global Report on Antimicrobial Resistance Surveillance, it is estimated that mortality due to resistant bacteria will ascend to 10 million cases per year worldwide. These facts highlight the importance of developing low-cost and readily accessible detection methods of drug-resistant ESBLs bacteria to prevent their spread and promote accurate and fast diagnosis. Bacterial detection is commonly done using molecular diagnostic techniques, where PCR stands out for its high performance. However, this technique requires specialized equipment not available everywhere, is time-consuming, and has a high cost. Loop-Mediated Isothermal Amplification (LAMP) is an alternative technique that works at a constant temperature, significantly decreasing the equipment cost. It yields double-stranded DNA of several lengths with repetitions of the target DNA sequence as a product. Although positive and negative results from LAMP can be discriminated by colorimetry, fluorescence, and turbidity, there is still a large room for improvement in the point-of-care implementation. DNA origami is a technique that allows the formation of 3D nanometric structures by folding a large single-stranded DNA (scaffold) into a determined shape with the help of short DNA sequences (staples), which hybridize with the scaffold. This research aimed to generate DNA origami structures using LAMP products as scaffolds to improve the sensitivity to detect ESBLs in point-of-care diagnosis. For this study, the coding sequence of the CTM-X-15 ESBL of E. coli was used to generate the LAMP products. The set of LAMP primers were designed using PrimerExplorerV5. As a result, a target sequence of 200 nucleotides from CTM-X-15 ESBL was obtained. Afterward, eight different DNA origami structures were designed using the target sequence in the SDCadnano and analyzed with CanDo to evaluate the stability of the 3D structures. The designs were constructed minimizing the total number of staples to reduce costs and complexity for point-of-care applications. After analyzing the DNA origami designs, two structures were selected. The first one was a zig-zag flat structure, while the second one was a wall-like shape. Given the sequence repetitions in the scaffold sequence, both were able to be assembled with only 6 different staples each one, ranging between 18 to 80 nucleotides. Simulations of both structures were performed using scaffolds of different sizes yielding stable structures in all the cases. The generation of the LAMP products were tested by colorimetry and electrophoresis. The formation of the DNA structures was analyzed using electrophoresis and colorimetry. The modeling of novel detection methods through bioinformatics tools allows reliable control and prediction of results. To our knowledge, this is the first study that uses LAMP products and DNA-origami in combination to delect ESBL-producing bacterial strains, which represent a promising methodology for diagnosis in the point-of-care.

Keywords: beta-lactamases, antibiotic resistance, DNA origami, isothermal amplification, LAMP technique, molecular diagnosis

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Authors: Khaled M. Khleifat

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Pseudomonas aeruginosa was isolated from infected burn patients and characterized by standard biochemical tests. The in vitro copper uptake was compared between this isolated pathogenic strain and two non-pathogenic control strains of Gram-positive bacteria Bacillusthuringiensis strain Israelisas well as Gram-negative bacteria Enterobacter aerogenes. Maximum copper uptake of 470 ppm/g biomass was obtained by P. aeruginosa strain, while the control strains B. thuringiensis and Enterobacter aerogenes had copper uptake of 350 and 383 ppm/g biomass, respectively. However, the lowest copper uptake (60 ppm/g biomass) was observed with another control the saprophytic strain Pseudomonas (Shewanella) putrefaciens. A further investigation regarding the effect of copper toxicity on bacterial growth, gave an MIC score of 600 ppm for P. aeruginosa strain compared to 460 and 300 ppm for the two Gram positive and Gram negative control strains, respectively. In tandem with these in vitro findings, blood analysis on burn patients infected with P. aeruginosa has indicated a selective decrease of copper (hypocupremia) and ceruloplasmin plasma levels. The iron metabolism was also affected by this copper deprivation leading to a similar decrease in plasma levels of PCV, iron, total iron-binding capacity, and transferrin. All these hematological changes were significantly different (P < 0.05) from the matched group of non-infected burn patients. The observed hypocupremia in infected burn patients was attributed to demanding scavenger ability by P. aeruginosa strain for the copper of plasma.

Keywords: Pseudomonas aeruginosa, hypocupremia, correlation, PCV

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Authors: Thayse Marques Passos, Brid Quilty, Mary Pryce

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The interest in developing alternative techniques to obtain safe water, free from pathogens and hazardous substances, is growing in recent times. The photodynamic inactivation of microorganisms (PDI) is a promising ecologically-friendly and multi-target approach for water disinfection. It uses visible light as an energy source combined with a photosensitiser (PS) to transfer energy/electrons to a substrate or molecular oxygen generating reactive oxygen species, which cause cidal effects towards cells. PDI has mainly been used in clinical studies and investigations on its application to disinfect water is relatively recent. The majority of studies use planktonic cells. However, in their natural environments, bacteria quite often do not occur as freely suspended cells (planktonic) but in cell aggregates that are either freely floating or attached to surfaces as biofilms. Microbes can form aggregates and biofilms as a strategy to protect them from environmental stress. As aggregates, bacteria have a better metabolic function, they communicate more efficiently, and they are more resistant to biocide compounds than their planktonic forms. Among the bacteria that are able to form aggregates are members of the genus Pseudomonas, they are a very diverse group widely distributed in the environment. Pseudomonas species can form aggregates/biofilms in water and can cause particular problems in water distribution systems. The aim of this study was to evaluate the effectiveness of photodynamic inactivation in killing a range of planktonic cells including Escherichia coli DSM 1103, Staphylococcus aureus DSM 799, Shigella sonnei DSM 5570, Salmonella enterica and Pseudomonas putida DSM 6125, and aggregating cells of Pseudomonas fluorescens DSM 50090, Pseudomonas aeruginosa PAO1. The experiments were performed in glass Petri dishes, containing the bacterial suspension and the photosensitiser, irradiated with a multi-LED (wavelengths 430nm and 660nm) for different time intervals. The responses of the cells were monitored using the pour plate technique and confocal microscopy. The study showed that bacteria belonging to Pseudomonads group tend to be more tolerant to PDI. While E. coli, S. aureus, S. sonnei and S. enterica required a dosage ranging from 39.47 J/cm2 to 59.21 J/cm2 for a 5 log reduction, Pseudomonads needed a dosage ranging from 78.94 to 118.42 J/cm2, a higher dose being required when the cells aggregated.

Keywords: bacterial aggregation, photoinactivation, Pseudomonads, water disinfection

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Authors: Vaibhav D. Bhatt, Anju P. Kunjadia, D. S. Nauriyal, Bhumika J. Joshi, Chaitanya G. Joshi

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Metagenomic analysis of milk samples collected from local cattle breed, kankrej (Bos indicus), Gir (Bos indicus) and Crossbred (Bos indicus X Bos taurus) cattle harbouring subclinical mastitis was carried out by next-generation sequencing (NGS) 454 GS-FLX technology. Around 56 different species including members of Enterobacteriales, Pseudomonadales, Bacillales and Lactobacillales with varying abundance were detected in infected milk. The interesting presence of bacteriophages against Staphylococcus aureus, Escherichia coli, Enterobacter and Yersinia species were observed, especially Enterobacteria and E. coli phages (0∙32%) in Kankrej, Enterobacteria and Staphylococcus phages (1∙05%) in Gir and Staphylococcus phages (2∙32%) in crossbred cattle. NGS findings suggest that phages may be involved in imparting natural resistance of the cattle against pathogens. Further infected milk samples were subjected for bacterial isolation. Fourteen different isolates were identified, and DNA was extracted. Genes (Tet-K, Msr-A, and Mec-A) providing antibiotic resistance to the bacteria were screened by Polymerase Chain Reaction and results were validated with traditional antibiotic assay. Total 3 bacteriophages were isolated from nearby environment of the cattle farm. The efficacy of phages was checked against multi-drug resistant bacteria, identified by PCR. In-vivo study was carried out for phage therapy in mammary glands of female rats “Wister albino”. Mammary glands were infused with MDR isolates for 3 consecutive days. Recovery was observed in infected rats after intramammary infusion of sterile phage suspension. From day 4th onwards, level of C-reactive protein was significant increases up to day 12th . However, significant reduction was observed between days 12th to 18th post treatment. Bacteriophages have significant potential as antibacterial agents and their ability to replicate exponentially within their hosts and their specificity, make them ideal candidates for more sustainable mastitis control.

Keywords: bacteriophages, c-reactive protein, mastitis, metagenomic analysis

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Authors: Mrunalini Sonne

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Aquaculture uses a variety of broad spectrum antibiotics to manage and prevent a variety of diseases without understanding their mechanisms of action. This has led to water pollution in the modern world. The necessity for alternate control measures against bacterial illnesses in the aquaculture sector is highlighted by issues brought on by antibiotic-resistant bacteria and the dearth of effective control strategies. Bacteriophages (phages) have shown promise as therapeutic agents for the efficient management of bacterial infections in aquaculture. In the current study, a variety of investigations were conducted to determine if utilizing lytic phages to reduce Aeromonas spp. infection in fish aquaculture was appropriate. Motile Aeromonas septicaemia is a fish disease that has caused financial harm to the aquaculture sector. Currently, the production of aquaculture depends significantly on antibiotics, which adds to the worldwide problem of the rise of bacteria that are resistant to medicines and resistance genes. To decrease the usage of antibiotics in aquaculture systems, it is crucial to create efficient antibiotic substitutes. Bacteriophages are capable of acting as a natural antagonist, mostly because of their great specificity, capacity for self-replication, and ability to quickly eradicate dangerous bacteria. There is a need for research that goes beyond just isolating and characterising lytic bacteriophages to examine their morphology, stability, and efficacy in various environmental conditions. Bacteriophage (or phage) therapy is a promising technique to control dangerous microbes in farmed fish. More phage therapy research in aquaculture is required in order to effectively employ phage treatment to lessen infection in fish brought on by Aeromonas.

Keywords: aquaculture, bacteriophages, fish, freshwater

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Authors: Sonja Djilas, Aleksandra Velićanski, Dragoljub Cvetković, Siniša Markov, Eva Lončar, Vesna Tumbas Šaponjac, Milica Vinčić

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Due to high content of bioactive compounds, sour cherry possesses antioxidant and antimicrobial activity. Additionally, waste material from industrial processing of sour cherry is also a good source of bioactive compounds. The aim of this study was to screen the antimicrobial activity and determine the minimal inhibitory (MIC) and minimal bactericidal concentrations (MBC) of sour cherry pomace extract. Tested strains were Gram-negative bacteria (Escherichia coli ATCC 25922, Salmonella typhimurium ATCC 14028 and wild isolates Escherichia coli and Salmonella sp.), Gram-positive bacteria (Staphylococcus aureus ATCC 11632, Bacillus cereus ATCC 10876 and wild isolates Staphylococcus saprophyticus and Bacillus sp.) and yeasts (Saccharomyces cerevisiae 112, Hefebank Weihenstephan and Candida albicans ATCC 10231). Antimicrobial activity was tested by disc-diffusion method and agar-well diffusion method. MIC and MBC were determined by microdilution method. Screening tests showed that Gram-negative bacteria were resistant to tested extract, with exception of Salmonella typhimurium and Salmonella sp. for which only zones of reduced growth appeared. However, Gram-positive bacteria were more sensitive where the highest clear zones appeared with 100 µl of extract applied. There was no activity against tested yeasts. MIC and MBC values were in the range 3.125-37.5 mg/ml and 6.25-100 mg/ml, respectively. The most susceptible strain was Staphylococcus aureus while the most resistant was Bacillus sp. where MBC was not found in tested concentration range. Sour cherry pomace possesses high antibacterial potential, which indicates that this waste material is a promising source of bioactive compounds and could be used as a functional food ingredient.

Keywords: antimicrobial activity, sour cherry, pomace, bioactive compounds

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Authors: Sweta Pandey, Samridhi Dhyani, Susmita Chaudhuri

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Klebsiella pneumoniae causes a wide range of infections, including urinary tract infections, sepsis, bacteremia, pneumonia, and liver abscesses. The emergence of multi-drug resistance in this bacterium led to a major setback for clinical management. WHO also endorsed a need for finding alternative therapy to antibiotics for the treatment of these infections. Development of vaccines and passive antibody therapy has been proven as a potent alternative to antibiotics in the case of MDR, XDR, and PDR Klebsiella infections. Siderophore receptors have been demonstrated to be overexpressed for the internalization of iron siderophore complexes during infections in most Gram-negative bacteria. For the present study, immune response to siderophore receptors to establish this protein as a potential immunogen for the development of therapeutic leads was explored. Clinical strains of Klebsiella pneumoniae were grown in iron-deficient conditions, and the iron-regulated outer membrane proteins were extracted and characterized through mass spectrometry for specific identification. The gene for identified protein was cloned in pET- 28a vector and expressed in E. coli. The native protein and the recombinant protein were isolated and purified and used as antigens for the generation of immune response in BALB/c mice. The native protein of Klebsiella pneumoniae grown in iron-deficient conditions was identified as FepA (Ferrienterobactin receptor) and other siderophore receptors. This 80 kDa protein generated an immune response in BALB/c mice. The antiserum from mice after subsequent booster doses was collected and showed binding with FepA protein in western blot and phagocytic uptake of the K. pneumoniae in the presence antiserum from immunized mice also observed from the animal studies after bacterial challenge post immunisation in mice have shown bacterial clearance. The antiserum from mice showed binding and clearance of the Klebsiella pneumoniae bacteria in vitro and in vivo. These antigens used for generating an active immune response in mice can further be used for therapeutic monoclonal antibody development against Klebsiella pneumoniae infections.

Keywords: antiserum, FepA, Klebsiella pneumoniae, multi drug resistance, siderophore receptor

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Authors: Nura Brimo, Dilek Cokeliler Serdaroglu, Tansel Uyar, Busra Uysal, Elif Bahar Cakici, Miris Dikmen, Zerrin Canturk

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Background: A combination of antibiotics, including metronidazole (MET), ciprofloxacin (CIP), and minocycline (MINO), has been demonstrated to disinfect bacteria in necrotic teeth before regenerative processes. It has been presented clinically that antibiotic pastes may drive to possible stem cell death and difficulties in removing from the canal system, which can limit the regenerative procedure. This study was designed to (1) synthesize nanofibrous webs containing various concentrations of different medicaments (triple, double, and calcium hydroxide,Ca(OH)2), and (2) coat thiselectrospun fibrous gutta-percha (GP) cones. Methods: Poly(vinylpyrrolidone) (PVP)-based electrospun fibrous webs were processed with low medicaments concentrations. Scanning Electron Microscopy (SEM), Energy Dispersive X-Ray Spectroscopy (EDX), and X-Ray Photoelectron Spectroscopy (XPS) were carried out to investigate fiber morphology, antibiotic incorporation, and characterized GP-coated fibrous webs, respectively. The chemical and physical properties of dentine were carried out via Fourier Transform Infrared Spectroscopy (FTIR) and Nano-SEM, respectively. The antimicrobial properties of the different fibrous webs were assessed against various bacteria by direct nanofiber/bacteria contact. Cytocompatibility was measured by applying the MTT method. Results: The mean fiber diameter of the experiment groups of medicament-containing fibers ranged in the nm scale and was significantly smaller than PVP fibers. EDX analysis confirmed the presence of medicaments in the nanofibers. XPS analysis presented a complete coating of the fibers with GPs; FTIR and Nano-SEM showed no chemical and physical configuration of intracanal medicaments on the dentine surface. Meanwhile, nanofibrous webs led to a significant reduction in the percentage of viable bacteria compared with the negative control and PVP. Conclusion: Our findings suggest that TA-NFs, DA-NFs, and Cₐ(OH)₂)-NFs coated GP cones have significant potential in eliminating intracanal bacteria, cell-friendly behavior, and clinical usage features.

Keywords: drug delivery, drug carrier, electrospinning, nano/microfibers, regenerative endodontic, morphology

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Authors: Khaled Khleifat, Muayyad Abboud, Amjad Khleifat, Humodi Saeed

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In this study, Pseudomonas aeruginosa was isolated from infected burn patients and characterized by standard biochemical tests. The in vitro copper uptake was compared between this isolated pathogenic strain and two non-pathogenic control strains of Gram positive bacteria Bacillusthuringiensis strain Israelisas well as Gram negative bacteria Enterobacter aerogenes. Maximum copper uptake of 470 ppm/g biomass was obtained by P. aeruginosa strain, while the control strains B. thuringiensis andEnterobacter aerogenes had copper uptake of 350 and 383 ppm/g biomass, respectively. However, the lowest copper uptake (60 ppm/g biomass) was observed with another control the saprophytic strain Pseudomonas (Shewanella) putrefaciens. A further investigation regarding the effect of copper toxicity on bacterial growth, gave an MIC score of 600 ppm for P. aeruginosa strain compared to 460 and 300 ppm for the two Gram positive and Gram negative control strains, respectively. In tandem with these in vitro findings, blood analysis on burn patients infected with P. aeruginosa has indicated a selective decrease of copper (hypocupremia) and ceruloplasmin plasma levels. The iron metabolism was also affected by this copper deprivation leading to a similar decrease in plasma levels of PCV, iron, total iron binding capacity, and transferrin. All these hematological changes were significantly different (P < 0.05) from the matched group of non-infected burn patients. The observed hypocupremia in infected burn patients was attributed to demanding scavenger ability by P. aeruginosa strain for the copper of plasma.

Keywords: pseudomonas, Cu uptake, burn patients, biosorption

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Authors: E. Bartkiene, M. Ruzauskas, V. Lele, P. Zavistanaviciute, J. Bernatoniene, V. Jakstas, L. Ivanauskas, D. Zadeike, D. Klupsaite, P. Viskelis, J. Bendoraitiene, V. Navikaite-Snipaitiene, G. Juodeikiene

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In this study, antimicrobial nutraceuticals; gummy candies (GC) from bovine colostrum (BC), essential oils (EOs), probiotic lactic acid bacteria (PLAB), and their combinations, were developed. For antimicrobial GC preparation, heteropolysaccharide (agar) was used. The antimicrobial properties of EOs (Eugenia caryophyllata, Thymus vulgaris, Citrus reticulata L., Citrus paradisi L.), BC, L. paracasei LUHS244, L. plantarum LUHS135, and their combinations against pathogenic bacteria strains (Streptococcus mutans, Enterococcus faecalis, Staphylococcus aureus, Salmonella enterica, Escherichia coli, Proteus mirabilis, and Pseudomonas aeruginosa) were evaluated. The highest antimicrobial properties by EO’s (Eugenia caryophyllata and Thymus vulgaris) were established. The optimal ingredients composition for antimicrobial GC preparation was established, which incorporate the BC fermented with L. paracasei LUHS244 in combination with Thymus vulgaris or Eugenia caryophyllata. These ingredients showed high inhibition properties of all tested pathogenic strains (except Pseudomonas aeruginosa). Antimicrobial GC formula consisting of thyme EO (up to 0.2%) and fermented BC (up to 3%), and for taste masking, mandarin or grapefruit EOs (up to 0.2%) was used. Developed GC high overall acceptability and antimicrobial properties, thus, antimicrobial GC could be a preferred form of nutraceuticals. This study was fulfilled with the support of the LSMU-KTU joint project.

Keywords: antimicrobial activity, bovine colostrum, essential oil, gummy candy, probiotic

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Authors: Asfa Hussain, Chee Tang, Mien Nguyen

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Introduction: The safe usage of medications is dependent on clear, well-documented prescribing. Medical drug charts should be regularly checked to ensure that they are fit for purpose. Aims: The purpose of this study was to evaluate whether the Isabel Hospice drug charts were effective or prone to medical errors. The aim was to create a comprehensive palliative care drug chart in line with medico-legal guidelines and to minimise drug administration and prescription errors. Methodology: 50 medical drug charts were audited from March to April 2020, to assess whether they complied with medico-legal guidelines, in a hospice within East of England. Meetings were held with the larger multi-disciplinary team (MDT), including the pharmacists, nursing staff and doctors, to raise awareness of the issue. A preliminary drug chart was created, using the input from the wider MDT. The chart was revised and trialled over 15 times, and each time feedback from the MDT was incorporated into the subsequent template. In the midst of the COVID-19 pandemic in September 2020, the finalised drug chart was trialled. 50 new palliative drug charts were re-audited, to evaluate the changes made. Results: Prescribing and administration errors were high prior to the implementation of the new chart. This improved significantly after introducing the new drug charts, therefore improving patient safety and care. The percentage of inadequately documented allergies went down from 66% to 20% and incorrect oxygen prescription from 40% to 16%. The prescription drug-drug interactions decreased by 30%. Conclusion: It is vital to have clear standardised drug charts, in line with medico-legal standards, to allow ease of prescription and administration of medications and ensure optimum patient-centred care. This closed loop audit demonstrated significant improvement in documentation and prevention of possible fatal drug errors and interactions.

Keywords: palliative care, drug chart, medication errors, drug-drug interactions, COVID-19, patient safety

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Authors: Sandeep Vasaikar, Lary Obi

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Rapid and discriminative genotyping methods are useful for determining the clonality of the isolates in nosocomial or household outbreaks. Multilocus sequence typing (MLST) is a nucleotide sequence-based approach for characterising bacterial isolates. The genetic diversity and the clinical relevance of the drug-resistant Klebsiella isolates from Mthatha are largely unknown. For this reason, prospective, experimental study of the molecular epidemiology of Klebsiella isolates from patients being treated in Mthatha over a three-year period was analysed. Methodology: PCR amplification and sequencing of the drug-resistance-associated genes, and multilocus sequence typing (MLST) using 7 housekeeping genes mdh, pgi, infB, FusAR, phoE, gapA and rpoB were conducted. A total of 32 isolates were analysed. Results: The percentages of multidrug-resistant (MDR), extensively drug-resistance (XDR) and pandrug-resistant (PDR) isolates were; MDR 65.6 % (21) and XDR and PDR with 0 % each. In this study, K. pneumoniae was 19/32 (59.4 %). MLST results showed 22 sequence types (STs) were identified, which were further separated by Maximum Parsimony into 10 clonal complexes and 12 singletons. The most dominant group was Klebsiella pneumoniae with 23/32 (71.8 %) isolates, Klebsiella oxytoca as a second group with 2/32 (6.25 %) isolates, and a single (3.1 %) K. varricola as a third group while 6 isolates were of unknown sequences. Conclusions/significance: A phylogenetic analysis of the concatenated sequences of the 7 housekeeping genes showed that strains of K. pneumoniae form a distinct lineage within the genus Klebsiella, with K. oxytoca and K. varricola its nearest phylogenetic neighbours. With the analysis of 7 genes were determined 1 K. variicola, which was mistakenly identified as K. pneumoniae by phenotypic methods. Two misidentifications of K. oxytoca were found when phenotypic methods were used. No significant differences were observed between ESBL blaCTX-M, blaTEM and blaSHV groups in the distribution of Sequence types (STs) or Clonal complexes (CCs).

Keywords: phylogenetic analysis, phylogeny, klebsiella phylogenetic, klebsiella

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Authors: Wilailak Siripornadulsil, Siriyanapat Tasaku, Jutamas Buahorm, Surasak Siripornadulsil

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The objectives of this study were to isolate LAB from various sources, dietary supplement, Thai traditional fermented food, and freshwater fish and to characterize their potential as probiotic cultures. Out of 1,558 isolates, 730 were identified as LAB based on isolation on MRS agar supplemented with a bromocresol purple indicator and CaCO3 and gram-positive, catalase and oxidase negative characteristics. Eight isolates showed the potential probiotic properties including tolerance to acid, bile salt and heat, proteolytic, amylolytic and lipolytic activities and oxalate-degrading capability. They all showed the antimicrobial activity against some Gram-negative and Gram-positive pathogenic bacteria. Based on 16S rDNA sequence analysis, they were identified as Enterococcus faecalis BT2 and MG30, Leconostoc mesenteroides SW64 and Pediococcus pentosaceous BD33, CF32, NP6, PS34 and SW5. The health beneficial effects and food safety will be further investigated and developed as a probiotic or protective culture used in Nile tilapia belly flap meat fermentation.

Keywords: probiotic, lactic acid bacteria, pathogen, protective culture

Procedia PDF Downloads 369

Authors: Norhan H. Mahdally, Bathini Thissera Riham A. ElShiekh, Noha M. Elhosseiny, Mona T. Kashef, Ali M. El Halawany, Mostafa E. Rateb, Ahmed S. Attia

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Multidrug-resistant (MDR) pathogens pose a global threat to healthcare settings. The exhaustion of the current antibiotic arsenal and the scarcity of new antimicrobials in the pipeline aggravate this threat and necessitate a prompt and effective response. This study focused on two major pathogens that can cause serious infections: carbapenem-resistant Acinetobacter baumannii (CRAB) and methicillin-resistant Staphylococcus aureus (MRSA). Multiple soil isolates were collected from several locations throughout Egypt and screened for their conventional and non-conventional antimicrobial activities against MDR pathogens. One isolate exhibited potent antimicrobial activity and was subjected to multiple rounds of fractionation. After fermentation and bio-guided fractionation, we identified pure microbial secondary metabolites with two scaffolds that exhibited promising effects against CRAB and MRSA. Scaling up and chemical synthesis of derivatives of the identified metabolite resulted in obtaining a more potent derivative, which we designated as 2HP. Cytotoxicity studies indicated that 2HP is well-tolerated by human cells. Ongoing work is focusing on formulating the new compound into a nano-formulation to enhance its delivery. Also, to have a better idea about how this compound works, a proteomic approach is currently underway. Our findings suggest that 2HP is a potential new broad-spectrum antimicrobial agent. Further studies are needed to confirm these findings and to develop 2HP into a safe and effective treatment for MDR infections.

Keywords: broad-spectrum antimicrobials, carbapenem-resistant acinetobacter baumannii, drug discovery, methicillin-resistant staphylococcus aureus, multidrug-resistant, natural products

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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

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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: Zohreh Bayat, Dariush Minai-Tehrani

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Pseudomonas aeruginosa is a Gram-negative, rode shape and aerobic bacterium that has shown to be resistance to many antibiotics. This resistance makes the bacterium very harmful in some diseases. It can also generate diseases in any part of the gastrointestinal tract from oropharynx to rectum. P. aeruginosa has become an important cause of infection, especially in patients with compromised host defense mechanisms. One of the most important reasons that make P. aeruginosa an emerging opportunistic pathogen in patients is its ability to use various compounds as carbon sources. Lipase is an enzyme that catalyzes the hydrolysis of lipids. Most lipases act at a specific position on the glycerol backbone of lipid substrate. Some lipases are expressed and secreted by pathogenic organisms during the infection. Muscarinic antagonist used as an antispasmodic and in urinary incontinence. The drug has little effect on glandular secretion or the cardiovascular system. It does have some local anesthetic properties and is used in gastrointestinal, biliary, and urinary tract spasms. Aim: In this study the inhibitory effect of a muscarinic antagonist on lipase of P. aeruginosa was investigated. Methods: P. aeruginosa was cultured in minimal salt medium with 1% olive oil as carbon source. The cells were harvested and the supernatant, which contained lipase, was used for enzyme assay. Results: Our results showed that the drug can inhibit P. aeruginosa lipase by competitive manner. In the presence of different concentrations of the drug, the Vmax (2 mmol/min/mg protein) of enzyme did not change, while the Km raised by increasing the drug concentration. The Ki (inhibition constant) and IC50 (the half maximal inhibitory concentration) value of drug was estimated to be about 30 uM and 60 uM which determined that the drug binds to enzyme with high affinity. Maximum activity of the enzyme was observed at pH 8 in the absence and presence of muscarinic antagonist, respectively. The maximum activity of lipase was observed at 600C and the enzyme became inactive at 900C. Conclusion: The muscarinic antagonist drug could inhibit lipase of P. aeruginosa and changed the kinetic parameters of the enzyme. The drug binded to enzyme with high affinity and did not chang the optimum pH of the enzyme. Temperature did not affect the binding of drug to musmuscarinic antagonist.

Keywords: Pseudomonas aeruginosa, drug, enzyme, inhibition

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Authors: M. H. Pazandeh, M. Doudi, S. Barahimi, L. Rahimzadeh Torabi

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The use of antibiotics is essential in reducing the occurrence of adverse effects and inhibiting the emergence of antibiotic resistance in microbial populations. The necessity for a novel methodology concerning local administration of antibiotics has arisen, with particular focus on dealing with localized infections prompted by bacterial colonization of medical devices or implant materials. Bioactive glasses (BG) are extensively employed in the field of regenerative medicine, encompassing a diverse range of materials utilized for drug delivery systems. In the present investigation, various drug carriers for imipenem and tetracycline, namely single systems BG/SnO2, BG/NiO with varying proportions of metal oxide, and nanocomposite BG/SnO2/NiO, were synthesized through the sol-gel technique. The antibacterial efficacy of the synthesized samples was assessed through the utilization of the disk diffusion method with the aim of neutralizing Staphylococcus aureus as the bacterial model. The current study involved the examination of the bioactivity of two samples, namely BG10SnO2/10NiO and BG20SnO2, which were chosen based on their heightened bacterial inactivation properties. This evaluation entailed the employment of two techniques: the measurement of the pH of simulated body fluid (SBF) solution and the analysis of the sample tablets through X-ray diffraction (XRD), scanning electron microscopy (SEM), and Fourier transform infrared (FTIR) spectroscopy. The sample tablets were submerged in SBF for varying durations of 7, 14, and 28 days. The bioactivity of the composite bioactive glass sample was assessed through characterization of alterations in its surface morphology, structure, and chemical composition. This evaluation was performed using scanning electron microscopy (SEM), Fourier-transform infrared (FTIR) spectroscopy, and X-ray diffraction spectroscopy. Subsequently, the sample was immersed in simulated liquids to simulate its behavior in biological environments. The specific body fat percentage (SBF) was assessed over a 28-day period. The confirmation of the formation of a hydroxyapatite surface layer serves as a distinct indicator of bioactivity. The infusion of antibiotics into the composite bioactive glass specimen was done separately, and then the release kinetics of tetracycline and imipenem were tested in simulated body fluid (SBF). Antimicrobial effectiveness against various bacterial strains have been proven in numerous instances using both melt and sol-gel techniques to create multiple bioactive glass compositions. An elevated concentration of calcium ions within a solution has been observed to cause an increase in the pH level. In aqueous suspensions, bioactive glass particles manifest a significant antimicrobial impact. The composite bioactive glass specimen exhibits a gradual and uninterrupted release, which is highly desirable for a drug delivery system over a span of 72 hours. The reduction in absorption, which signals the loss of a portion of the antibiotic during the loading process from the initial phosphate-buffered saline solution, indicates the successful bonding of the two antibiotics to the surfaces of the bioactive glass samples. The sample denoted as BG/10SnO2/10NiO exhibits a higher loading of particles compared to the sample designated as BG/20SnO2 in the context of bioactive glass. The enriched sample demonstrates a heightened bactericidal impact on the bacteria under investigation while concurrently preserving its antibacterial characteristics. Tailored bioactive glass that incorporates hydroxyapatite, with a regulated and efficient release of drugs targeting bacterial infections, holds promise as a potential framework for bone implant scaffolds following rigorous clinical evaluation, thereby establishing potential future biomedical uses. During the modification process, the introduction of metal oxides into bioactive glass resulted in improved antibacterial characteristics, particularly in the composite bioactive glass sample that displayed the highest level of efficiency.

Keywords: antibacterial, bioactive glasses, implant infections, multi drug resistant

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Authors: Kubilay Kurtulus Bastas

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Bacterial diseases are very destructive and cause economic losses on pears. Promising plant extracts for the management of plant diseases are environmentally safe, long-lasting and extracts of certain plants contain alkaloids, tannins, quinones, coumarins, phenolic compounds, and phytoalexins. In this study, bacteria were isolated from different parts of pear exhibiting characteristic symptoms of bacterial diseases from the Central Anatolia, Turkey. Pathogenic bacteria were identified by morphological, physiological, biochemical and molecular methods as fire blight (Erwinia amylovora (39%)), bacterial blossom blast and blister bark (Pseudomonas syringae pv. syringae (22%)), crown gall (Rhizobium radiobacter (1%)) from different pear cultivars, and determined virulence levels of the pathogens with pathogenicity tests. The air-dried 25 plant material was ground into fine powder and extraction was performed at room temperature by maceration with 80% (v/v) methanol/distilled water. The minimum inhibitory concentration (MIC) values were determined by using modified disc diffusion method at five different concentrations and streptomycin sulphate was used as control chemical. Bacterial suspensions were prepared as 108 CFU ml⁻¹ densities and 100 µl bacterial suspensions were spread to TSA medium. Antimicrobial activity was evaluated by measuring the inhibition zones in reference to the test organisms. Among the tested plants, Origanum vulgare, Hedera helix, Satureja hortensis, Rhus coriaria, Eucalyptus globulus, Rosmarinus officinalis, Ocimum basilicum, Salvia officinalis, Cuminum cyminum and Thymus vulgaris showed a good antibacterial activity and they inhibited the growth of the pathogens with inhibition zone diameter ranging from 7 to 27 mm at 20% (w/v) in absolute methanol in vitro conditions. In vivo, the highest efficacy was determined as 27% on reducing tumor formation of R. radiobacter, and 48% and 41% on reducing shoot blight of E. amylovora and P. s. pv. syringae on pear seedlings, respectively. Obtaining data indicated that some plant extracts may be used against the bacterial diseases on pome fruits within sustainable and organic management programs.

Keywords: bacteria, eco-friendly management, organic, pear, plant extract

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Authors: Deepika Bhaskar, Neena Wadehra, Megha Gulati, Aruna Narula, R. Vishnu, Gunjan Katyal

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With drug resistance becoming widespread in Plasmodium falciparum infections, development of the alternative drugs is the desired strategy for prevention and cure of malaria. Three drug targets were selected to screen promising drug molecules from the GSK library of around 14000 molecules. Using an in silico structure-based drug designing approach, the differences in binding energies of the substrate and inhibitor were exploited between target sites of parasite and human to design a drug molecule against Plasmodium. The docking studies have shown several promising molecules from GSK library with more effective binding as compared to the already known inhibitors for the drug targets. Though stronger interaction has been shown by several molecules as compare to reference, few molecules have shown the potential as drug candidates though in vitro studies are required to validate the results.

Keywords: plasmodium, malaria, drug targets, in silico studies

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Authors: C. T. Eze, I. N. E. Onwurah

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An important hydrogeologic problem in areas of high faults formations is high environmental health hazard occasioned by microbial and heavy metals contamination of ground waters. Consequently, we examined the microbial load and heavy metals concentration of hospital wastewater discharged into the environment at Park Lane General Hospital Enugu Urban, Nigeria. The microbial counts, characteristics and frequency of occurrences of the isolated microorganisms were determined by cultural, morphological and biochemical characteristics using established procedure while the varying concentrations of the identified heavy metals were determined using the spectrophotometric method. The microbiological analyses showed a mean total aerobic bacteria counts from 13.7 ± 0.65 × 107 to 22.8 ± 1.14 ×1010 CFU/ml, mean total anaerobic bacteria counts from 6.0 ± 1.6 × 103 to 1.7 ± 0.41 ×104 CFU/ml and mean total fungal counts from 0 ± 0 to 2.3 ± 0.16 × 105 CFU/ml. The isolated micro-organisms which included both pathogenic and non-pathogenic organisms were Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa, Salmonella typhi, Bacillus subtilis, Proteus vulgaris, Klesbsiella pneumonia and bacteriodes sp. The only fungal isolate was Candida albican. The heavy metals identified in the leachate were Arsenic, Cadmium, Lead, Mercury and Chromium and their concentrations ranged from 0.003 ± 0.00082 to 0.14 ± 0.0082 mg/l. These values were above WHO permissible limits while others fall within the limits. Therefore, hospital waste water can pose the environmental health risk when not properly treated before discharge, especially in geologic formations with high fault formations.

Keywords: bacterial isolates, fungal isolates, heavy metals, hospital wastewater, microbial counts

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Authors: M. E. Abalaka, S. Y. Daniyan, S. O. Adeyemo, D. Damisa

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Gold nanoparticles (AuNPs) have gained increasing interest in recent times. This is greatly due to their special features, which include unusual optical and electronic properties, high stability and biological compatibility, controllable morphology and size dispersion, and easy surface functionalization. In typical synthesis, AuNPs were produced by reduction of gold salt AuCl4 in an appropriate solvent. A stabilizing agent was added to prevent the particles from aggregating. The antibacterial activity of different sizes of gold nanoparticles was investigated against Staphylococcus aureus, Salmonella typhi and Pseudomonas pneumonia using the disk diffusion method in a Müeller–Hinton Agar. The Au-NPs were effective against all bacteria tested. That the Au-NPs were successfully synthesized in suspension and were used to study the antibacterial activity of the two medicinal plants against some bacterial pathogens suggests that Au-NPs can be employed as an effective bacteria inhibitor and may be an effective tool in medical field. The study clearly showed that the Au-NPs exhibiting inhibition towards the tested pathogenic bacteria in vitro could have the same effects in vivo and thus may be useful in the medical field if well researched into.

Keywords: gold nanoparticles, Gomphrena celesioides, Prunus amygdalus, pathogens

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Authors: Suyeon Kwon, Ik Joong Kang, Wang Bingjie

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Chitosan is a polymer that is usually produced from N-deacetylation of chitin. It is emerging as a promising biocompatible polymer that is harmless to humans. For the reason that many merits such as good adsorptive, biodegradability, many researches are being done on the chitosan for drug delivery system. Drug delivery system (DDS) has been developed for the control of drug. It makes the drug can be delivered effectively and safely into the targeted human body. The drug used in this work is hydrocortisone that is used in Rheumatism, skin diseases, allergy treatment. In this work, hydrocortisone was used to make allergic rhinitis medicine. Our study focuses on drug delivery through the nasal mucosa by using hydrocortisone impregnated chitosan nanoshells. This study has performed an investigation in order to establish the optimal conditions, changing concentration, quantity of hydrocortisone. DLS, SEM, TEM, FT-IR, UV spectrum were used to analyze the manufactured chitosan-hydrocortisone silver nanoshell and silver nanoshell, whose function as drug carriers. This study has performed an investigation on new drug carriers and delivery routes for hydrocortisone. Various methods of manufacturing chitosan-hydrocortisone nanoshells were attempted in order to establish the optimal condition. As a result, the average size of chitosan-hydrocortisone silver nanoshell is about 80 nm. So, chitosan-hydrocortisone silver nanoshell is suitable as drug carriers because optimal size of drug carrier in human body is less than 120 nm. UV spectrum of Chitosan-hydrocortisone silver nanoshell shows the characteristic peak of silver nanoshell at 420 nm. Likewise, the average size of chitosan-hydrocortisone silver nanoshell is about 100nm. It is also suitable for drug carrier in human body. Also, multi-layered silver shell over chitosan nanoshells induced the red-shift of absorption peak and increased the intensity of absorption peak. The resultant chitosan–silver nanocomposites (or nanoshells) exhibited the absorption peak around 430nm attributed to silvershell formation. i.e. the absorption peak was red-shifted by ca. 40 nm in reference to 390 nm of silver nanoshells.

Keywords: chitosan, drug delivery, hydrocortisone, rhinitis, nanoshell

Procedia PDF Downloads 251

Authors: Sara Ataei, Molouk Hadjibabaie, Shirinsadat Badri, Amirhossein Moslehi, Iman Karimzadeh, Ardeshir Ghavamzadeh

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Purpose: To assess the pattern and probable risk factors for moderate and major drug–drug interactions in a referral hematology-oncology ward in Iran. Methods: All patients admitted to hematology–oncology ward of Dr. Shariati Hospital during a 6-month period and received at least two anti-cancer or non-anti-cancer medications simultaneously were included. All being scheduled anti-cancer and non-anti-cancer medications both prescribed and administered during ward stay were considered for drug–drug interaction screening by Lexi-Interact On- Desktop software. Results: One hundred and eighty-five drug–drug interactions with moderate or major severity were detected from 83 patients. Most of drug–drug interactions (69.73 %) were classified as pharmacokinetics. Fluconazole (25.95 %) was the most commonly offending medication in drug–drug interactions. Interaction of sulfamethoxazole-trimethoprim with fluconazole was the most common drug–drug interaction (27.27 %). Vincristine with imatinib was the only identified interaction between two anti-cancer agents. The number of administered medications during ward stay was considered as an independent risk factor for developing a drug–drug interaction. Conclusions: Potential moderate or major drug–drug interactions occur frequently in patients with hematological malignancies or related diseases. Performing larger standard studies are required to assess the real clinical and economical effects of drug–drug interactions on patients with hematological and non-hematological malignancies.

Keywords: drug–drug interactions, hematology–oncology ward, hematological malignancies

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Authors: Neelam Taneja, Abhishek Mewara, Ajay Kumar

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Multidrug resistant shigellae have emerged as a therapeutic challenge in India. At our 2000 bed tertiary care referral centre in Chandigarh, North India, which caters to a large population of 7 neighboring states, antibiotic resistance in Shigella is being constantly monitored. Shigellae are isolated from 3 to 5% of all stool samples. In 1990 nalidixic acid was the drug of choice as 82%, and 63% of shigellae were resistant to ampicillin and cotrimoxazole respectively. Nalidixic acid resistance emerged in 1992 and rapidly increased from 6% during 1994-98 to 86% by the turn of 21st century. In the 1990s, the WHO recommended ciprofloxacin as the drug of choice for empiric treatment of shigellosis in view of the existing high level resistance to agents like chloramphenicol, ampicillin, cotrimoxazole and nalidixic acid. First resistance to ciprofloxacin in S. flexneri at our centre appeared in 2000 and rapidly rose to 46% in 2007 (MIC>4mg/L). In between we had an outbreak of ciprofloxacin resistant S.dysenteriae serotype 1 in 2003. Therapeutic failures with ciprofloxacin occurred with both ciprofloxacin-resistant S. dysenteriae and ciprofloxacin-resistant S. flexneri. The severity of illness was more with ciprofloxacin-resistant strains. Till 2000, elsewhere in the world ciprofloxacin resistance in S. flexneri was sporadic and uncommon, though resistance to co-trimoxazole and ampicillin was common and in some areas resistance to nalidixic acid had also emerged. Fluoroquinolones due to extensive use and misuse for many other illnesses in our region are thus no longer the preferred group of drugs for managing shigellosis in India. WHO presently recommends ceftriaxone and azithromycin as alternative drugs to fluoroquinolone-resistant shigellae, however, overreliance on this group of drugs also seems to soon become questionable considering the emerging cephalosporin-resistant shigellae. We found 15.1% of S. flexneri isolates collected over a period of 9 years (2000-2009) resistant to at least one of the third-generation cephalosporins (ceftriaxone/cefotaxime). The first isolate showing ceftriaxone resistance was obtained in 2001, and we have observed an increase in number of isolates resistant to third generation cephalosporins in S. flexneri 2005 onwards. This situation has now become a therapeutic challenge in our region. The MIC values for Shigella isolates revealed a worrisome rise for ceftriaxone (MIC90:12 mg/L) and cefepime (MIC90:8 mg/L). MIC values for S. dysenteriae remained below 1 mg/L for ceftriaxone, however for cefepime, the MIC90 has raised to 4 mg/L. These infections caused by ceftriaxone-resistant S. flexneri isolates were successfully treated by azithromycin at our center. Most worrisome development in the present has been the emergence of DSA(Decreased susceptibility to azithromycin) which surfaced in 2001 and has increased from 4.3% till 2011 to 34% thereafter. We suspect plasmid-mediated resistance as we detected qnrS1-positive Shigella for the first time from the Indian subcontinent in 2 strains from 2010, indicating a relatively new appearance of this PMQR determinant among Shigella in India. This calls for a continuous and strong surveillance of antibiotic resistance across the country. The prevention of shigellosis by developing cost-effective vaccines is desirable as it will substantially reduce the morbidity associated with diarrhoea in the country

Keywords: Shigella, antimicrobial, resistance, India

Procedia PDF Downloads 224

Authors: Adela Diepoltova, Klara Konecna, Ondrej Jandourek, Petr Nachtigal

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A loss of control over antibiotic-resistant pathogens has become a global issue due to severe and often untreatable infections. This state is reflected in complicated treatment, health costs, and higher mortality. All these factors emphasize the urgent need for the discovery and development of new anti-infectives. One of the most common pathogens mentioned in the phenomenon of antibiotic resistance are bacteria of the genus Staphylococcus. These bacterial agents have developed several mechanisms against the effect of antibiotics. One of them is biofilm formation. In staphylococci, biofilms are associated with infections such as endocarditis, osteomyelitis, catheter-related bloodstream infections, etc. To author's best knowledge, no validated and standardized methodology evaluating candidate compound activity against staphylococcal biofilms exists. However, a variety of protocols for in vitro drug activity testing has been suggested, yet there are often fundamental differences. Based on our experience, a key methodological step that leads to credible results is to form a robust biofilm with appropriate attributes such as firm adherence to the substrate, a complex arrangement in layers, and the presence of extracellular polysaccharide matrix. At first, for the purpose of drug antibiofilm activity evaluation, the focus was put on various conditions (supplementation of cultivation media by human plasma/fetal bovine serum, shaking mode, the density of initial inoculum) that should lead to reproducible and robust in vitro staphylococcal biofilm formation in microtiter plate model. Three model staphylococcal reference strains were included in the study: Staphylococcus aureus (ATCC 29213), methicillin-resistant Staphylococcus aureus (ATCC 43300), and Staphylococcus epidermidis (ATCC 35983). The total biofilm biomass was quantified using the Christensen method with crystal violet, and results obtained from at least three independent experiments were statistically processed. Attention was also paid to the viability of the biofilm-forming staphylococcal cells and the presence of extracellular polysaccharide matrix. The conditions that led to robust biofilm biomass formation with attributes for biofilms mentioned above were then applied by introducing an alternative method analogous to the commercially available test system, the Calgary Biofilm Device. In this test system, biofilms are formed on pegs that are incorporated into the lid of the microtiter plate. This system provides several advantages (in situ detection and quantification of biofilm microbial cells that have retained their viability after drug exposure). Based on our preliminary studies, it was found that the attention to the peg surface and substrate on which the bacterial biofilms are formed should also be paid to. Therefore, further steps leading to the optimization were introduced. The surface of pegs was coated by human plasma, fetal bovine serum, and L-polylysine. Subsequently, the willingness of bacteria to adhere and form biofilm was monitored. In conclusion, suitable conditions were revealed, leading to the formation of reproducible, robust staphylococcal biofilms in vitro for the microtiter model and the system analogous to the Calgary biofilm device, as well. The robustness and typical slime texture could be detected visually. Likewise, an analysis by confocal laser scanning microscopy revealed a complex three-dimensional arrangement of biofilm forming organisms surrounded by an extracellular polysaccharide matrix.

Keywords: anti-biofilm drug activity screening, in vitro biofilm formation, microtiter plate model, the Calgary biofilm device, staphylococcal infections, substrate modification, surface coating

Procedia PDF Downloads 144

Authors: Marcelus U. Ajonina, Deodata B. Ngonga, Kenric B. Ware, Carine K. Nfor

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Background: Lack of knowledge of rational use of antimalarial drugs among dispensers is a serious problem, especially in areas of intense transmission, thus increasing the risk of resistance and adverse drug reactions. This study was aimed at assessing the knowledge of malaria as well as perception and dispensing practices of antimalarials among vendors in Buea community. Methods: A community-based cross-sectional survey of a random sample of 140 drug vendors living within the Buea community was conducted between March and June 2017. A questionnaire was designed to obtain information from drug vendors on the general knowledge of malaria as well as dispensing practices. Data were analyzed using SPSS Statistics 20.0 and were considered significant at p ≤ 0.05. Results: Knowledge of malaria symptoms, transmission, and prevention was reasonable among 55.8% (77) of the respondents. Only 33.6% (47) of the respondents could attribute the cause of malaria to protozoan of genus Plasmodium species. Of the 140 vendors, 115 (82.7%) prescribe antimalarial drugs. The knowledge of the national protocol was malaria case management among dispensers was 35.0%. Vendors in hospital/community pharmacies were 2.4 times (OR = 3.14, 95% CI: 4.14 - 8.74, p < 0.001) more knowledgeable about malaria treatment protocol than those of in drugstores. The prevalence of self-prescription of antimalarials was 39.3%. Self-prescription was significantly higher in drugstores than hospital/community pharmacies (p=0.004). In all, 56 (40.6%) of vendors showed good practices regarding antimalarial drug dispensing with the majority (51.7%) from community pharmacies (OR=2.27,95% CI: 1.13-4.56). Conclusion: Findings reveal moderate knowledge of malaria but poor prescription and dispensing practices of antimalarial drugs among vendors, thus indicating a need for routine monitoring and evaluation to prevent the emergence of resistant strains to current efficacious antimalarials.

Keywords: antimalarials, drug retail outlets, dispensing, drug resistance, prescription

Procedia PDF Downloads 124

Authors: Jarel Elgin Tolentino, Arby Denise Nera, Mary Rose Roco, Angela Vianca Aspa, Nikko Beltran, Else Dapat

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Drug resistance by cells has been the problem in the medical field for decades now. The use of medicinal plants as a source of creating powerful drugs has been nowadays recognized worldwide to treat such resistant diseases. In the present study, the potential for Diospyros philippinensis A. DC. to inhibit growth of both bacteria and cancer cell line was conducted. The leaf crude extracts were screened for the presence of phytochemicals and examined for potential bioactivities by employing several assays like Kirby-Bauer disc diffusion method, DPPH (2,2-diphenyl-1-picryl-hydrazyl-hydrate) assay and MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) tetrazolium assay for the antibacterial, antioxidant and cytotoxic activities of the extract, respectively. Phytochemical test results of the extracts revealed the presence of alkaloids, flavonoids, saponins, phenols, quinones, cardiac glycosides, phlobatannins, carbohydrate, cardenolides and proteins. The leaf extracts were found to exhibit antibacterial activity against gram-positive bacteria, high antioxidant activity (99.22% ± 0.005) but did not show any sign of cytotoxicity towards HCT116 (ATCC CCL-247). The study therefore concludes that D. philippinensis A. DC. leaf extract can be a source of antibacterial and chemopreventive agents. This claim may be used as basis for future investigation.

Keywords: bioassay, medicinal plants, plant crude extracts, phytochemical screening

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Authors: Asif Khaliq, Sayeeda A. Sayed

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The drug information centers provide drug related information to the requesters that include physicians, pharmacist, nurses and other allied health care professionals. The International Pharmacist Federation (FIP) describes basic functions of a drug and poison information centers as drug evaluation, therapeutic counseling, pharmaceutical advice, research, pharmaco-vigilence and toxicology. Continuous advancement in the field of medicine has expanded the medical literature, which has increased demand of a drug and poison information center for the guidance, support and facilitation of physicians. The objective of the study is to determine the need of drug and poison information centers in public and private hospitals of Karachi, Pakistan. A cross sectional study was conducted during July 2013 to April 2014 using a self-administered, multi-itemed questionnaire. Non Probability Convenient sampling was used to select the study participants. A total of 307 physicians from public and private hospitals of Karachi participated in the study. The need for 24/7 Drug and poison information center was highlighted by 92 % of physicians and 67% physicians suggested opening a drug information center at the hospital. It was reported that 70% physicians take at least 15 minutes for searching the information about the drug while managing a case. Regarding the poisoning case management, 52% physicians complaint about the unavailability of medicines in hospitals; and mentioned the importance of medicines for safe and timely management of patients. Although 73% physicians attended continued medical education (CME) sessions, 92 % physicians insisted on the need of 24/7 Drug and poison information center. The scarcity of organized channel for obtaining the information about drug and poisons is one of the most crucial problems for healthcare workers in Pakistan. The drug and poison information center is an advisory body that assists health care professional and patients in provision of appropriate drug and hazardous substance information. Drug and poison information center is one of the integral needs for running an effective health care system. Provision of a 24 /7 drug information centers with specialized staff offer multiple benefits to the hospitals while reducing treatment delays, addressing awareness gaps of all stakeholders and ensuring provision of quality health care.

Keywords: drug and poison information centers, Pakistan, physicians, public and private hospitals

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Authors: Nelson Rotimi Osungbemiro, Sanni Rafiu Olugbenga, Abayomi Olufemi Olajuyigbe

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Ninety Fingerlings of Clarias gariepinus were exposed to the pathogenic Flavobacterium columnare a Gram Negative bacteria responsible for high mortality in fish pond raised young fish (fries and fingerlings) of Clarias sp. in Southwestern Nigeria. After feeding with 40% crude protein pelletized fish feed for 5 days, the fishes were divided into two groups, one group was treated with extracts from Calendula officinalis flowers, while the second group was not treated (control). The results indicated that, at day 5, colony formation had been manifesting and at day 7, skin lesion occurred and at the 8th day, first mortality of fish occurred, and this continued steadily on the 9th-12th day when all the fishes were dead. Whereas, in the group that was treated with Calendula sp., no single mortality was recorded. This research shows that plant extract from Calendula flowers is an effective antimicrobial agent against the virulent pathogenic Flavobacterium columnare disease.

Keywords: antimicrobial, Flavobacterium columnare, Clarias gariepinus, fish

Procedia PDF Downloads 593