The completion of Escherichia coli K1 genome has identified several genomic islands that are present in meningitis-causing E. coli RS218 but absent in the non-pathogenic E. coli MG1655. In this study, the role of various genomic islands in E. coli K1 interactions with intestinal epithelial cells (Caco-2) and kidney epithelial cells (MA104) was determined. Using association assays, invasion assays, and intracellular survival assays, the findings revealed that the genomic island deletion mutants of RS218 related to P fimbriae, S fimbriae, F17-like fimbriae, non-fimbrial adhesins, Hek and hemagglutinin, protein secretion system (T1SS for hemolysin; T2SS; T5SS for antigen 43), Iro system and hmu system), invasins (CNF1, IbeA), toxins (α-hemolysin), K1 capsule biosynthesis, metabolism (d-serine catabolism, dihydroxyacetone, glycerol, and glyoxylate metabolism), prophage genes, showed reduced interactions with both cell types. Next, we determined the role of various genomic islands in E. coli K1 resistance to serum. When exposed to the normal human serum, the viability of the genomic island deletion mutants related to adhesins such as S fimbriae, P fimbriae, F17-like fimbriae, non-fimbrial adhesins, Hek and hemagglutinin, antigen 43 and T5SS for antigen 43, T2SS, and T1SS for hemolysin, Iro system and hmu system, prophage genes, metabolism (sugar metabolism and d-serine catabolism), K1 capsule biosynthesis, and invasins such as CNF1 was affected, suggesting their role in bacteremia. The characterization of these genomic islands should reveal mechanisms of E. coli K1 pathogenicity that could be of value as therapeutic targets.
Acinetobacter baumannii is a Gram-negative nosocomial pathogen of importance due to its uncanny ability to acquire resistance to most antimicrobials. These include carbapenems, which are the drugs of choice for treating A. baumannii infections, and polymyxins, the drugs of last resort. Whole genome sequencing was performed on two clinical carbapenem-resistant A. baumannii AC29 and AC30 strains which had an indistinguishable ApaI pulsotype but different susceptibilities to polymyxin. Both genomes consisted of an approximately 3.8 Mbp circular chromosome each and several plasmids. AC29 (susceptible to polymyxin) and AC30 (resistant to polymyxin) belonged to the ST195 lineage and are phylogenetically clustered under the International Clone II (IC-II) group. An AbaR4-type resistance island (RI) interrupted the comM gene in the chromosomes of both strains and contained the bla OXA-23 carbapenemase gene and determinants for tetracycline and streptomycin resistance. AC29 harbored another copy of bla OXA-23 in a large (~74 kb) conjugative plasmid, pAC29b, but this gene was absent in a similar plasmid (pAC30c) found in AC30. A 7 kb Tn1548::armA RI which encodes determinants for aminoglycoside and macrolide resistance, is chromosomally-located in AC29 but found in a 16 kb plasmid in AC30, pAC30b. Analysis of known determinants for polymyxin resistance in AC30 showed mutations in the pmrA gene encoding the response regulator of the two-component pmrAB signal transduction system as well as in the lpxD, lpxC, and lpsB genes that encode enzymes involved in the biosynthesis of lipopolysaccharide (LPS). Experimental evidence indicated that impairment of LPS along with overexpression of pmrAB may have contributed to the development of polymyxin resistance in AC30. Cloning of a novel variant of the bla AmpC gene from AC29 and AC30, and its subsequent expression in E. coli also indicated its likely function as an extended-spectrum cephalosporinase.
Cell-cell communication is also known as quorum sensing (QS) that happens in the bacterial cells with the aim to regulate their genes expression in response to increased cell density. In this study, a bacterium (L8A) isolated from dental plaque biofilm was identified as Citrobacter amalonaticus by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry (MS). Its N-acylhomoserine-lactone (AHL) production was screened by using two types of AHL biosensors namely Chromobacterium violaceum CV026 and Escherichia coli [pSB401]. Citrobacter amalonaticus strain L8A was identified and confirmed producing numerous types of AHL namely N-butyryl-L-homoserine lactone (C4-HSL), N-hexanoyl-L-homoserine lactone (C6-HSL), N-octanoyl-L-homoserine lactone (C8-HSL) and N-hexadecanoyl-L-homoserine lactone (C16-HSL). We performed the whole genome sequence analysis of this oral isolate where its genome sequence reveals the presence of QS signal synthase gene and our work will pave the ways to study the function of the related QS genes in this bacterium.
The study explored on the commonly available weed plant Commelina nudiflora which has potential in-vitro antioxidant and antimicrobial activity. The different polar solvents such as ethanol, chloroform, dichloromethane, hexane and aqueous were used for the soxhlet extraction. The extracts were identified pharmacologically as important bioactive compounds and their potential free radical scavenging activities, and antimicrobial properties were studied. C. nudiflora extracts were monitored on their in-vitro antioxidant ability by DPPH and ABTS radical scavenging assay. Aqueous extract shows significant free radical scavenging activity of 63.4 mg/GAE and 49.10 mg/g in DPPH and ABTS respectively. Furthermore, the aqueous crude extract was used in antibacterial studies, which shows the highest inhibitory activity against Pseudomonas aeruginosa, Escherichia coli and Salmonella typhi. Among all the extracts, aqueous extract of C. nudiflora has significant control over free radical scavenging activity and inhibition of the growth of food pathogenic bacteria. Also, the aqueous extract contains abundance of phenolics and flavonoids higher than other extracts. This study explored weed plant C. nudiflora as a potential source of antioxidant and antibacterial efficacy and identified various therapeutic value bioactive compounds from GC-MS analysis.
Myriad proteobacteria use N-acyl homoserine lactone (AHL) molecules as quorum sensing (QS) signals to regulate different physiological functions, including virulence, antibiotic production, and biofilm formation. Many of these proteobacteria possess LuxI/LuxR system as the QS mechanism. Recently, we reported the 3.89 Mb genome of Acinetobacter sp. strain GG2. In this work, the genome of this long chain AHL-producing bacterium was unravelled which led to the molecular characterization of luxI homologue, designated as aciI. This 552 bp gene was cloned and overexpressed in Escherichia coli BL21(DE3). The purified protein was ∼20.5 kDa and is highly similar to several autoinducer proteins of LuxI family among Acinetobacter species. To verify the AHL synthesis activity of this protein, high-resolution liquid chromatography-mass spectrometry analysis revealed the production of 3-oxo-dodecanoyl-homoserine lactone and 3-hydroxy-dodecanoyl-homoserine lactone from induced E. coli harboring the recombinant AciI. Our data show for the first time, the cloning and characterization of the luxI homologue from Acinetobacter sp. strain GG2, and confirmation of its AHLs production. These data are of great significance as the annotated genome of strain GG2 has provided a valuable insight in the study of autoinducer molecules and its roles in QS mechanism of the bacterium.
Systems metabolic engineering and in silico analyses are necessary to study gene knockout candidate for enhanced succinic acid production by Escherichia coli. Metabolically engineered E. coli has been reported to produce succinate from glucose and glycerol. However, investigation on in silico deletion of ptsG/b1101 gene in E. coli from glycerol using minimization of metabolic adjustment algorithm with the OptFlux software platform has not yet been elucidated. Herein we report what is to our knowledge the first direct predicted increase in succinate production following in silico deletion of the ptsG gene in E. coli GEM from glycerol with the OptFlux software platform. The result indicates that the deletion of this gene in E. coli GEM predicts increased succinate production that is 20% higher than the wild-type control model. Hence, the mutant model maintained a growth rate that is 77% of the wild-type parent model. It was established that knocking out of the ptsG/b1101 gene in E. coli using glucose as substrate enhanced succinate production, but the exact mechanism of this effect is still obscure. This study informs other studies that the deletion of ptsG/b1101 gene in E. coli GEM predicted increased succinate production, enabling a model-driven experimental inquiry and/or novel biological discovery on the underground metabolic role of this gene in E. coli central metabolism in relation to increasing succinate production when glycerol is the substrate.
Extended-spectrum β-lactamase (ESBL) producing Enterobacteriaceae are often susceptible in vitro to β-lactam/β-lactamase inhibitor (BLBLI) combination antibiotics, but their use has been limited by concerns of clinical inefficacy. We aimed to compare outcomes between patients treated with BLBLIs and carbapenems for bloodstream infection (BSI) caused by cefotaxime non-susceptible (likely ESBL- or AmpC β-lactamase-producing) Escherichia coli and Klebsiella pneumoniae.
Thirty-five veterinary isolates of Salmonella enteritidis were characterized by their susceptibility to 10 antimicrobial agents and by their plasmid profiles on agarose gel electrophoresis. All were susceptible to carbenicillin, chloramphenicol and nalidixic acid but 89% were resistant to tetracycline. When examined, 91% of the isolates harboured plasmids, with sizes ranging from 9.8 to 60 MDa. However, it was only possible to associate the presence of plasmids with tetracycline resistance; plasmids occurring in 90% of the tetracycline-resistant isolates. In conjugation experiments, with Escherichia coli K12 Nal(r) as recipient, the tetracycline resistance in three selected S. enteritidis isolates was observed to transfer at frequencies of 3.0×10(-3) to 1.0×10(-2)/donor cell. The concomitant transfer of a 56-MDa or 60-MDa plasmid in these three S. enteritidis isolates was also detected.
In a case study involving preliminary investigations for the feasibility of a beach resort complex in the west coast of Sarawak, Malaysia, since the acceptable quality of recreational water quality had not been specified, existing international standards and practices were reviewed to arrive at acceptable microbiological and physicochemical parametric levels. Water samples were collected on a weekly basis in the proposed beach complex at Tanjong Batu Coastal Reserve and also along the nearby Sungei Batang Kemena and its estuary. It was ascertained that the swimming water quality was acceptable in terms of faecal Coliforms, temperature, pH and dissolved oxygen. However E. Coli counts did indicate a potential risk of 1.68% for gastrointestinal illness and the ubiquitous presence of faecal Streptococci pointed to recent pollution of human or animal origin. Besides, grease and oil contents exceeded the WHO Guideline values. It was hence recommended that an appropriate sanitation or pollution survey should be carried out in the adjacent coastal catchment area and the beach. Also routine water sampling should be undertaken. Such action will help to pinpoint sources of pollution and lead to antipollution measures, thus helping to upgrade swimming water quality and establish swimming water quality standards.
The pattern of antibiotic resistance amongst gram-negative bacteria (GNB) in paediatric units, which have heavy empirical usage of broad-spectrum antibiotics, was studied prospectively over a 6-month period. A total of 200 consecutive, non-duplicate gram-negative isolates were obtained from 109 patients admitted to intensive care and oncology units in two hospitals. The commonest isolates were Klebsiella spp (36.5 per cent) and Pseudomonas (20.0 per cent). The isolates showed lower susceptibility rates to the third-generation cephalosporins (47-62 per cent) compared with cefepime (91 per cent), imipenem (90 per cent) and ciprofloxacin (99 per cent). Fifty-four (52.8 per cent) Klebsiella and Escherichia coli isolates were determined to be extended-spectrum beta-lactamase (ESBL) producing strains. Antibiotics found to be effective against ESBL-producers were imipenem and ciprofloxacin. The high resistance rate amongst GNB to third-generation cephalosporins is a likely consequence of heavy empirical usage of this group of antibiotics. The carbapenems and quinolones remain useful agents in the management of patients admitted to these units.
Bacteriological isolation and identification were performed on 60 cloacal swabs and 15 aquarium water samples of pet red-eared sliders (Trachemys scripta elegans) obtained from aquarium shops in the Klang Valley, Central Peninsula Malaysia. The most common bacteria isolated was Aeromonas spp., which was present in both cloacal swabs (70%) and aquarium water (86.7%). Klebsiella spp. (50%), Escherichia coli (33.3%), Yersinia spp. (16.7%) and Salmonella spp. (15%) obtained form cloacal swabs were identified as pathogenic to both humans and animals. Salmonella spp. were isolated from both cloacal swabs and aquarium water. The Salmonella serotypes identified were S. tennessee, S. typhimurium, S. brezany, S. pomona, S. corvallis and S. schwarzengrund. Bacterial infections in humans associated with handling exotic pets directly or indirectly in contact with aquarium water have been described regularly, hence the zoonotic significance of owning a turtle infected with Salmonella spp. or any pathogenic bacteria therefore cannot be ignored.
Introduction:Staphylococcus aureus is a Gram-positive staphylococci that form biofilms. Bacteria that dwell in bio-films tend to be highly resistant towards the action of antibiotics. S. aureus is a main cause of infections in the oral cavity such as angular cheilitis, endodontic infections, osteomyelitis of the jaw, parotitis and oral mucositis. Previous studies reported that S. aureus also spread to the other parts of the body through the circulatory system, which may lead to chronic infections. Hence the search for new antibacterial agents remains high and needs urgent attention to treat this problem. Plants offer a rich source of antimicrobial agents and bioactive compounds. In this study, aque-ous oil palm leaf extracts (OPLE) has been used as an alternative antibacterial agent against oral infections mainly caused by Staphylococcus aureus. Many studies report the potential use of oil palm leaf extracts in treating bacterial infections such as Escherichia coli, Salmonella sp., Staphylococcus aureus (isolated from other part of the body), Pseudomonas aeruginosa and Bacillus sp. Although previous studies have documented the antimicrobial properties of oil palm leaf extracts, to date no study has been reported on the effect of oil palm leaf extract on oral microbes. Methods: The agar diffusion method, minimum inhibitory concentration (MIC) and minimal bactericidal concen-tration (MBC) assay were conducted in order to observe the antibacterial activity of aqueous oil palm leaf extract. The crystal violet assay was used to determine the anti-biofilm activity of the extracts. Chlorhexidine and deionised distilled water were used as the positive and negative control respectively. For agar diffusion method, the diameter of inhibition zone was measured. Results: The inhibition zone of the tested bacteria was observed between 0-20mm. The MIC and MBC assay were used to know the lowest concentrations of the extract that inhibit the growth and killed the tested bacteria respectively. The MIC and MBC values for the tested bacteria were observed between 0-7.813mg/mL. While for anti-biofilm assays, OPLE aqueous extract acts as a potent anti-biofilm agent with dual actions, pre-venting and eradicating the biofilm of the tested bacteria. Conclusion: In conclusion, the tested plant extracts could serve as alternative natural antibacterial and anti-biofilm agent against oral infections.
Biofouling on the membrane surface leads to performance deficiencies in membrane filtration. In this study, the application of ginger extract as a bio-based additive to enhance membrane antibiofouling properties was investigated. The extract was dispersed in a dimethyl acetamide (DMAc) solvent together with polyvinylidene fluoride (PVDF) to enhance biofouling resistance of the resulting membrane due to its antibiotic property. The concentrations of the ginger extract in the dope solution were varied in the range of 0-0.1 wt %. The antibacterial property of the resulting membranes was assessed using the Kirby Bauer disc diffusion method. The results show an inhibition zone formed around the PVDF/ginger membrane against Escherichia coli and Staphylococcus aureus demonstrating the efficacy of the residual ginger extract in the membrane matrix to impose the antibiofouling property. The addition of the ginger extract also enhanced the hydrophilicity in the membrane surface by lowering the contact angle from 93° to 85°, which was in good agreement with the increase in the pure water flux of up to 62%.
The evolution of cosmetic products results in the growing demands for cosmetics that are preservatives free. Plant essential oils were found to be a promising antimicrobial and also antioxidant agent. In this study, Cymbopogon citratus (lemongrass), Laurus nobilis (bay leaf) and Backhousia citriodora (lemon myrtle) essential oils were selected and evaluated for their antimicrobial properties. It was found that Laurus nobilis exhibited strong antimicrobial activity against the selected bacteria Streptococcus saprophyticus (ATCC 49619), Streptococcus aureus (ATCC 22923), Streptococcus pyogenes (ATCC 29436), Pseudomonas aeruginosa (ATCC 13048), Klebsiella pneumoniae (ATCC 700603), Escherichia coli (ATCC 22922) with MIC ranging between 7.8 ug/mL to 250 μg/mL. The antioxidant activity of selected essential oils was determined by antioxidant assays which were 1,1-Diphenyl-2-picrylhydrazyl assay (DPPH), determination of ferric reducing antioxidant power assay (FRAP) and β-Carotene/linoleic acid bleaching assay. Backhousia citriodora and Laurus nobilis showed the highest antioxidant activity.
n-Octanal and β-Selinene were identified to be the major components with peak area of 26.37 % and 13.92 % respectively in secondary metabolites analysis by Gas Chromatography-Mass Spectrometry (GCMS).
Demand for milk has increased in Malaysia due to the increased in awareness of healthy foods consumption.
Hence, research of milk is crucial to ensure that it is not contaminated with Escherichia coli. This study
evaluated the survival of Escherichia coli at different temperature and haemolysin activity of Escherichia
coli on blood agar. A total of 8 samples of raw fresh and pasteurized milk were collected from nearby farm
and market in Negeri Sembilan, Malaysia. After an overnight exposure to four different temperatures of
0
0C, 280C, 350C and 450C, the bacteriological test of milk was evaluated for the presence of Escherichia
coli. Overall, all raw fresh milk sampled exceeded the acceptable limit of bacterial count of 1 x 105 CFU/ml.
Raw fresh milk recorded the highest count at 35oC with 4.4 x 107 CFU/ml and the lowest at 0oC with 8.3 x
104 CFU/ml. The presence of Escherichia coli was detected in 7/20(35%) of the total raw fresh milk
samples. All pasteurized milk showed no presence of Escherichia coli due to the effectiveness of heat
treatment. Haemolysin test showed no haemolytic activity. Milk contaminated with Escherichia coli can
cause diarrheal, gastrointestinal diseases and urinary infection. Hence, it is important to study the survival
rate of Escherichia coli and its pathogenicity in milk to ensure public safety.
Engineering the CO2
-fixing enzyme ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) to improve photosynthesis
has long been sought. Rubisco large subunits (RbcL) are highly-conserved but because of certain undefined sequence
differences, plant Rubisco research cannot fully utilise the robust heterologous Escherichia coli expression system and its
GroEL folding machinery. Previously, a series of chimeric cyanobacteria Synechococcus elongatus Rubisco, incorporated
with sequences from the green alga Chlamydomonas reinhardtii, were expressed in E. coli; differences in RbcL sections
essential for holoenzyme formation were pinpointed. In this study, the remaining sections, presumably not crucial for
holoenzyme formation and also the small subunit (RbcS), are substituted to further ascertain the possible destabilising
effects of multiple section mutations. To that end, combinations of Synechococcus RbcL Sections 1 (residues 1-47), 2
(residues 48-97), 5 (residues 198-247) and 10 (residues 448-472), and RbcS, were swapped with collinear Chlamydomonas
sections and expressed in E. coli. Interestingly, only the chimera with Sections 1 and 2 together produces holoenzyme and
an interaction network of complementing amino acid changes is delineated by crystal structure analysis. Furthermore,
sequence-based analysis also highlighted possible GroEL binding site differences between the two RbcLs.
In the present study, gelatin-carboxymethyl cellulose blend film was synthesized, characterized and applied for the first time to preserve cherry tomatoes (Solanum lycopersicum var. cerasiforme) and grapes (Vitis vinifera). Gelatin (Gel) film forming solution was incorporated with carboxymethyl cellulose (CMC) at three volume per volume (Gel:CMC) ratios, namely 75:25, 50:50 and 25:75. CMC treatment has improved the transparency, tensile strength (TS), elongation at break (EAB), water vapor permeability and oxygen permeability of gelatin films. A pronounced effect was obtained for 25Gel:75CMC film. The TS and EAB values were increased from 25.98 MPa and 2.34% (100Gel:0CMC) to 37.54 MPa and 4.41% (25Gel:75CMC), respectively. A significant improvement in antimicrobial property of gelatin films against two food pathogens, namely Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus) was obtained in the presence of CMC. The effectiveness of gelatin-CMC blend films to extend the shelf life of agricultural products was evaluated in a 14-day preservation study. The gelatin-CMC films were successfully controlled the weight loss and browning index of the fruits up to 50.41% and 31.34%, respectively. Overall, gelatin-CMC film is an environmental friendly film for food preservation.
This research work was executed to determine chemical composition, anti-oxidant and anti-microbial potential of the essential oils extracted from the leaves and stem of Daphne mucronata Royle. From leaves and stem oils fifty-one different constituents were identified through GC/MS examination. The antioxidant potential evaluated through DPPH free radical scavenging activity and %-inhibition of peroxidation in linoleic acid system. The stem's essential oil showed the good antioxidant activity as compared to leaves essential oil. Results of Antimicrobial activity revealed that both stem and leaves oils showed strong activity against Candida albicans with large inhibition zone (22.2 ± 0.01, 18.9 ± 0.20 mm) and lowest MIC values (0.98 ± 0.005, 2.44 ± 0.002 mg/mL) respectively. Leaves essential was also active against Escherichia coli with inhibition zone of 8.88 ± 0.01 mm and MIC values of 11.2 ± 0.40 mg/mL. These results suggested that the plant's essential oils would be a potential cradle for the natural product based antimicrobial as well as antioxidant agents.
BACKGROUND: A series of 6, 6'-(1,4-phenylene)bis(4-(4-bromophenyl)pyrimidin-2-amine) derivatives has been synthesized by Claisen-Schmidt condensation and its chemical structures was confirmed by FT-IR, 1H/13C-NMR spectral and elemental analyses. The molecular docking study was carried out to find the interaction between active bis-pyrimidine compounds with CDK-8 protein. The in vitro antimicrobial potential of the synthesized compounds was determined against Gram-positive and Gram-negative bacterial species as well fungal species by tube dilution technique. Antimicrobial results indicated that compound 11y was found to be most potent one against E. coli (MICec = 0.67 µmol/mL) and C. albicans (MICca = 0.17 µmol/mL) and its activity was comparable to norfloxacin (MIC = 0.47 µmol/mL) and fluconazole (MIC = 0.50 µmol/mL), respectively.
CONCLUSION: Anticancer screening of the synthesized compounds using Sulforhodamine B (SRB) assay demonstrated that compounds 2y (IC50 = 0.01 µmol/mL) and 4y (IC50= 0.02 µmol/mL) have high antiproliferative potential against human colorectal carcinoma cancer cell line than the reference drug (5- fluorouracil) and these compounds also showed best dock score with better potency within the ATP binding pocket and may also be used lead for rational drug designing.
The electrospinning PAN nanofiber membrane (P-CN) was hydrolysed to convert carboxylic groups as reaction sites and covalently graft chitosan molecule. The chitosan derivatives with quaternary ammonium groups exerted greater efficiency against bacteria as compared to pure chitosan. Hence, the chitosan modified membrane (P-CS), can be functionalized with quaternary amine (i.e., glycidyl trimethyl ammonium chloride, GTMAC) to form quaternized chitosan nanofiber membrane (designated as P-HTCC) under various conditions (acidic, neutral, and alkaline). N-quaternized derivatives of chitosan modified membrane (N-HTCC) showed 72% and 60% degree of quaternization (DQ) under acidic and neutral conditions, respectively. Under alkaline condition, additional quaternization of N, O-HTCC via its amino and hydroxyl groups, has improved up to 90% DQ of the chitosan. The antibacterial activity of the quaternized chitosan modified membrane prepared from acetic acid medium is stronger than that prepared from water and alkaline media. Also, antibacterial activity of quaternized chitosan is stronger than chitosan modified membrane against E. coli. The microbiological assessments showed that the water-stable P-HTCC nanofiber membrane under modification in acidic medium exerted antibacterial activity up to 99.95% against E. coli. Therefore, the P-HTCC membrane exhibited high potential to be integrated into microfiltration membrane to effectively disinfect E. coli.