Home > Isolation and Molecular Characterization of Colicin/Microcin Producing Probiotic Escherichia Coli and its Therapeutic Utility Against Selected Gram Negative Pathogens
Isolation and Molecular Characterization of Colicin/Microcin Producing Probiotic Escherichia Coli and its Therapeutic Utility Against Selected Gram Negative Pathogens
Probiotics are living microbial adjuncts which has valuable effects on host body incuding improved food digestion, enhancing host‟s immune responses, modification of host associated microbial community and thus, improving the overall ambient environment. Since increase in multidrug resistance pathogens makes the standard treatments in-effective, probiotic have been developed sccessfully as alternate therapeutic drugs in recent era. Commercialized synthetic drugs (e.g. sulfa drugs) have also been interlinked with severe side effects to the hosts which have ultimately lemmatized their use. Bacteriocin (peptide antimicrobial) producing bacteria that can competitively exclude pathogens in the host, is an emrging approach towards novel therapeutics. These defensive proteins are released by 95% of bacterial species in their natural habitat to compete with one another for food and space. Purified bacteriocins of Lactobactericeae (lantibiotics) have been commercialized as prebiotics after successful experiments in laboratory animal. Bacteriocins of Gram negative bacteria (colicins and microcins) are also able to competitively exclude closely related bacterial species when fed to model organism and livestock. The current study was carried out to isolate commensal Gram negative strains that can hinder the growth of common pathogens by producing colicins and microcins against them. Commensal E.coli strains from cattle, sheep and humans were the choice of study since they were expected to better adapt to the host body when administrated as probiotics. Phenotypic inhibitory activity was determined, followed by genomic detection of various colicins, microcins and virulence determinants among the active isolates. Moreover, these strains were also evaluated for antibiotic sensitivity profile, pH tolerance, hemolytic activity and entero-invasive property. The E.coli strains were also sorted into phylogenetic groups of E.coli i.e. A0, A1, B1, B2, D1 and D2. From 300 fecal samples of cattle, sheep and human, 675 lactose fermenting Gram negative colonies were selected, out of which 513 were confirmed as E.coli based on biochemical characterization. For accuracy of further tests, 465 E.coli isolates (n=155/ sample) were evaluated in vitro for growth inhibition of five Gram negative species i.e. E.coli O157:H7, E.coli O26:H11, Salmonella enterica, Klebsiella pneumonia and Pseudomonas aeruginosa. Only 68 isolates (26.4% cattle, 25% sheep and 48.5% human) produced significant growth inhibition among which 41.1% inhibited E.coli O157:H7, 35.2% E.coli O26:H11, 30.7% S. enterica, 24.8% P. aeruginosa and 13% inhibited K. pneumonia. Genomic enumeration of colicin and microcin determinants of these antagonistic E.coli isolates revealed that each of the isolate produced at-least one colicin while the microcins were rarely found in commensal species. The highest prevailing colicin was col E6 (70.4%) followed by col Ib (66.3%), E4 (54.28%), E7 (49.9%), col J (42.9%), col M (35.2%) and col Ia (29.3%). Colicins like S4, D, A, E1, E3 and microcins including mH47, mV, mB17 and mC7 were prevalent at intermediate levels (< 25%) while col E2, 10 and mB17 were rarely found (< 5%) in E.coli isolates. Determinants for col B, K, 5 and mL were not perceived in any E.coli isolate. Phylogenetic grouping revealed that group B2 E.coli was more prevelant (i.e. 27.9%) among human samples followed by 13.2% in sheep and 11.7% in cattle while group D1 was the second most prevalent group i.e. 16%. Almost 13.2 % B1 E.coli, 11.6% A1 and 5.8% A0 E.coli were found among the antagonistic strains from all sources. Virulence gene determinants (i.e. Stx1, Stx2, Hlye, St and eaeA) were detected in 60.2% E.coli isolates while remaining 39.7% were non-pathogenic commensal isolates which were further confirmed by 16SrRNA sequencing. Antibiotic susceptibility profile, hemolysis assay, entero-invasive ability and pH tolerance of all non-pathogenic strains revealed that out of total E.coli strains, four cattle, two human and one sheep derived E.coli strains were the most suitable strains to be considered as probiotic species. However, further insight is suggested to unveil the effects of downstream processes and to discover eminent capsule packaging for these strains so that they can be easily fed to hosts.
منزلِ شوق کے رنگ ڈاکٹر نوید عاجزؔ ڈاکٹر فہد ملک کا پہلا تعارف انسانیت کے مسیحا کا ہے۔ لیکن وہ روح کی مسیحائی پر یقین رکھتے ہوئے شاعری کو مقدم گردانتے ہیں۔ سچائی، خلوص اور انسان دوستی کی صفات اُن کی شخصیت کی تعمیر میں کلیدی حیثیت رکھتی ہیں۔ چونکہ وہ شاعری کو شخصیت کے اظہار کا وسیلہ تصور کرتے ہیں، اس لیے ان کے کلام میں بھی مذکورہ صفات پوری آب و تاب کے ساتھ جلوہ گر نظر آتی ہیں۔ وہ کہتے ہیں: عداوتوں کو تیرگی، وفا کو روشنی کہا تمام عمر ہم نے تو جو سچ تھا بس وہی کہا غزل کی روایت کا ایک موضوع احباب کی بے وفائی کا ہے۔ یہ موضوع اس قدر آفاقیت کا حامل ہے کہ ہر دور و ہر دیار میں پنپتا ہوا مل جائے گا۔ہمارے معاشرتی نظام میں دولت کی چمک دمک انسان کے ضمیر کو سلا کر وقتی مفادات پر مبنی فیصلے لینے پر مجبور کر دیتی ہے لیکن بے وفائی کی کوئی توجیہہ قابلِ قبول نہیں۔ یہ موضوع شاعری کا سدا بہار حصہ ہے۔ بقول فہد ملک: ہم راہ دیکھتے رہے تھے جس کی عمر بھر وہ شہر میں جو آیا تو ہم سے نہیں ملا فہد کے اندر حساس انسان اُسے غور و فکر پر اکسائے رکھتا ہے، جس کی دلیل یہ ہے کہ اس کے ہاں فلسفیانہ مزاج کے اشعار ملتے ہیں۔ وجودی کرب انسان کا مقدر ہے۔ جو شخص دنیا میں آ گیا ہے وہ غم ہائے روزگار سے جان نہیں چھڑا سکتا۔ میرؔ و غالبؔ سے قاسمیؔ اور فرازؔ تک سبھی اس کرب کو محسوس کرتے اور پھر بیان کرتے نظر آتے ہیں۔ انسانی تخلیق کے حوالے سے فہد بھی اسی کرب سے گزرتے ہوئے لکھتا ہے: اس گردشِ جہان کا مارا ہوا وجود لے جائیں ہم کہاں بھلا ہارا ہوا وجود
E-Commerce in today’s world is playing an important role in global business. A quantitative and quantitative research was conducted with professional members, employees working in organization which are dealing with E-Business or online working via internet and their views gave an empirical and dynamic approach to make business efficient and effective. Connectivity with market to find best manufacture for goods and for manufacture to finds its buyer for their finished goods. Flow of information become faster and gathering of data become easier to make activities of business to get optimal results. Numbers of goods are available complications to finds goods are removed by complimentary goods. Supplies of goods at proper time in order to meet demand by consumer plays win to win game. Improved methods of technology like RFID make work easier and accessible information about goods to management. Manpower in forms of labor is reducing which leads to decrease in the cost of goods. Ordering cost saves time and gives a specified time both producers to make commodity and consumer for delivery of goods. E-Commerce with contrast of components of supply chain in the form of ware house, logistics, number of listing suppliers, number of listing items on one web in the form of website on one platform give dynamic image to businesses in the race of competitive world. Business transactions done with internet not only reduce time but also make documents or records for longer time able to send data from distant parts of the world. Through websites we can get completive price information and other relevant information for goods.
This electrochemical study of three different biologically active compounds has been conducted using cyclic voltammetry technique at gold electrode. Losartan Potassium is an antihypertensive drug, while Gemifloxacin is antibacterial and Clarithromycin is primarily bacteriostatic and also has antimicrobial effect. Cyclic voltammetric study has been conducted by using (0.04M) Britton Robinson Buffer as supporting electrolyte with different pH range. For Losartan Potassium pH range of B-R buffer was 8-11 while for Clarithromycin and Gemifloxacin B-R buffer pH range (2-6) has been selected according to the appropriate solubility of these pharmaceutical compounds. Voltammograms of all three biologically active compounds have been recorded at six different scan rates of 20, 100, 200, 300, 400 and 500mV/s. Different electrochemical parameters such as peak potential (Ep), peak current (Ip), transfer coefficient (α), number of electron (nα), diffusion coefficient (D), and heterogeneous rate constant (K0) were determined. Moreover, diagnostics tests have also been applied to define the electrochemical properties of these compounds. Results indicate that Losartan Potassium follows electrochemically irreversible reduction process with transfer of two electrons involving adsorption controlled process on gold electrode. However, electrochemical behavior of Gemifloxacin showed quasi reversible redox process with two electron transfer and on the electrode surface some adsorption complications have been observed. In case of Clarithromycin irreversible oxidation process with two electron transfer has been identified and electrode processes were shown to be diffusion controlled. These quantitative and qualitative investigations based on cyclic voltammetry technique demonstrate that this method is very reliable, sensitive and appropriate for the determination of electrochemical properties of different biological and pharmaceutical compounds using gold electrode. Moreover, this technique can also be used for quality control and pharmacokinetics studies of biologically active compounds.