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      Multi-drug resistant pathogenic bacteria in the gut of young children in Bangladesh

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          Abstract

          Background

          The gut of human harbors diverse commensal microbiota performing an array of beneficial role for the hosts. In the present study, the major commensal gut bacteria isolated by culturing methods from 15 children of moderate income families, aged between 10 and 24 months, were studied for their response to different antibiotics, and the molecular basis of drug resistance.

          Results

          Of 122 bacterial colonies primarily selected from Luria–Bertani agar, bacterial genera confirmed by analytical profile index (API) 20E ® system included Escherichia as the predominant (52%) organism, followed by Enterobacter (16%), Pseudomonas (12%), Klebsiella (6%), Pantoea (6%), Vibrio (3%), and Citrobacter (3%); while Aeromonas and Raoultella were identified as the infrequently occurring genera. An estimated 11 and 22% of the E. coli isolates carried virulence marker genes stx-2 and eae, respectively. Antimicrobial susceptibility assay revealed 78% of the gut bacteria to be multidrug resistant (MDR) with highest resistance to erythromycin (96%), followed by ampicillin (63%), tetracycline (59%), azithromycin (53%), sulfamethoxazole-trimethoprim (43%), cefixime (39%), and ceftriaxone (33%). PCR assay results revealed 56% of the gut bacteria to possess gene cassette Class 1 integron; while 8, 17.5 and 6% of the strains carried tetracycline resistance-related genes tetA, tetB, and tetD, respectively. The macrolide (erythromycin and azithromycin) resistance marker genes mphA, ereB, and ermB were found in 28, 3 and 5% of bacterial isolates, respectively; while 26, 12, 17, 32, 7, 4 and 3% of the MDR bacterial isolates carried the extended spectrum β-lactamase (ESBL)-related genes e.g., bla TEM, bla SHV, bla CMY-9, bla CTX-M1, bla CTX-M2, bla CMY-2 and bla OXA respectively. Majority of the MDR gut bacteria harbored large plasmids [e.g., 140 MDa (43%), 105 MDa (30%), 90 MDa (14%)] carrying invasion and related antibiotic resistance marker genes.

          Conclusions

          Our results suggest gut of young Bangladeshi children to be an important reservoir for multi-drug resistant pathogenic bacteria carrying ESBL related genes.

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          Most cited references33

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          Rapid procedure for detection and isolation of large and small plasmids.

          Procedures are described for the detection and isolation of plasmids of various sizes (2.6 to 350 megadaltons) that are harbored in species of Agrobacterium, Rhizobium, Escherichia, Salmonella, Erwinia, Pseudomonas, and Xanthomonas. The method utilized the molecular characteristics of covalently closed circular deoxyribonucleic acid (DNA) that is released from cells under conditions that denature chromosomal DNA by using alkaline sodium dodecyl sulfate (pH 12.6) at elevated temperatures. Proteins and cell debris were removed by extraction with phenol-chloroform. Under these conditions chromosomal DNA concentrations were reduced or eliminated. The clarified extract was used directly for electrophoretic analysis. These procedures also permitted the selective isolation of plasmid DNA that can be used directly in nick translation, restriction endonuclease analysis, transformation, and DNA cloning experiments.
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            Mechanisms of resistance in multiple-antibiotic-resistant Escherichia coli strains of human, animal, and food origins.

            Seventeen multiple-antibiotic-resistant nonpathogenic Escherichia coli strains of human, animal, and food origins showed a wide variety of antibiotic resistance genes, many of them carried by class 1 and class 2 integrons. Amino acid changes in MarR and mutations in marO were identified for 15 and 14 E. coli strains, respectively.
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              Gut Microbiota of Healthy and Malnourished Children in Bangladesh

              Poor health and malnutrition in preschool children are longstanding problems in Bangladesh. Gut microbiota plays a tremendous role in nutrient absorption and determining the state of health. In this study, metagenomic tool was employed to assess the gut microbiota composition of healthy and malnourished children. DNA was extracted from fecal samples of seven healthy and seven malnourished children (n = 14; age 2–3 years) were analyzed for the variable region of 16S rRNA genes by universal primer PCR followed by high-throughput 454 parallel sequencing to identify the bacterial phyla and genera. Our results reveal that the healthy children had a significantly higher number of operational taxonomic unit in their gut than that of the malnourished children (healthy vs. malnourished: 546 vs. 310). In malnourished children, bacterial population of the phyla Proteobacteria and Bacteroidetes accounted for 46 and 18%, respectively. Conversely, in healthy children, Proteobacteria and Bacteroidetes accounted for 5% and 44, respectively (p < 0.001). In malnourished children, the phylum Proteobacteria included pathogenic genera, namely Klebsiella and Escherichia, which were 174-fold and 9-fold higher, respectively, than their healthy counterpart. The predominance of potentially pathogenic Proteobacteria and minimal level of Bacteroidetes as commensal microbiota might be associated to the ill health of malnourished children in Bangladesh.
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                Author and article information

                Contributors
                smonira@icddrb.org
                syeda.shabnam@gmail.com
                imranali.mbo@gmail.com
                sadique004@gmail.com
                mbmita@gmail.com
                Zillur.rahman@icddrb.org
                nhalam@icddrb.org
                watanabe-haruo@iuhw.ac.jp
                9827001-10 , munirul@icddrb.org
                Journal
                Gut Pathog
                Gut Pathog
                Gut Pathogens
                BioMed Central (London )
                1757-4749
                20 April 2017
                20 April 2017
                2017
                : 9
                : 19
                Affiliations
                [1 ]ISNI 0000 0004 0600 7174, GRID grid.414142.6, , International Center for Diarrheal Disease Research, Bangladesh (icddr,b), ; 68 Shaheed Tajuddin Ahmed Sarani, Mohakhali, Dhaka, 1212 Bangladesh
                [2 ]ISNI 0000 0001 2220 1880, GRID grid.410795.e, , National Institute of Infectious Diseases, ; Tokyo, Japan
                Article
                170
                10.1186/s13099-017-0170-4
                5399343
                28439298
                0283f0d0-88b8-4263-b29e-7e222f4412e5
                © The Author(s) 2017

                Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License ( http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver ( http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.

                History
                : 9 January 2017
                : 10 April 2017
                Funding
                Funded by: icddr,b and NIID, Japan
                Categories
                Research
                Custom metadata
                © The Author(s) 2017

                Gastroenterology & Hepatology
                children,gut,microbiota,multidrug resistance,esbl related genes
                Gastroenterology & Hepatology
                children, gut, microbiota, multidrug resistance, esbl related genes

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