scholarly journals 549. First Report for Emergence of Chromosomal Borne Colistin Resistance Gene mcr-1 in a Clinical Acinetobacter Baumannii Isolates from India

2019 ◽  
Vol 6 (Supplement_2) ◽  
pp. S261-S262 ◽  
Author(s):  
MOHIBUR RAHMAN ◽  
Saheem Ahmad
Keyword(s):  
Acinetobacter Baumannii ◽  
Resistance Gene ◽  
Gel Electrophoresis ◽  
Southern Hybridization ◽  
Antibiotic Resistance Gene ◽  
Pulsed Field Gel Electrophoresis ◽  
Colistin Resistance ◽  
Tertiary Referral Hospital ◽  
Pulsed Field ◽  
First Case

Abstract Background Efficacy of Colistin the last line agent against infections due to multidrug-resistant (MDR) gram-negative pathogens, has been challenged when Liu et al. reported a plasmid-mediated gene, mcr-1, in 2015. Thereby this plasmid-borne mcr has been reported in bacterial isolates worldwide taken from humans, animals, farms, foods, and the environment. The present work invesitigate the mcr gene among clinical isolates of Acinetobacter Baumannii at our tertiary referral hospital of India. Methods The study was conducted at Sanjay Gandhi Post Graduate Institute of Medical Sciences, Lucknow, India. MIC values for 100 consecutive non-duplicate MDR isolates of Acinetobacter were checked for Colistin. PCR amplification of mcr gene was performed followed by sequencing of the amplicons. Clinical features of patients infected with mcr positive isolates were unveiled. Clonal relatedness of these isolates was investigated by Pulsed-field gel electrophoresis (PFGE). The mcr-1 localization was checked by conjugation followed by PFGE southern hybridization. Results 20/100 (20%) isolates were colistin resistant with having MIC Values of more than 8≤ µg/mL. The 20 colistin resistances isolates were PCR positive for mcr-1 and had been assigned EMBL/GeneBank nucleotide accession numbers MH730099-MH730118. Oher antibiotic resistance gene like ESBL, NDM-1, VIM, and 16s rRNA methyl transferases like Arm A, rmtC, rmt F were also found in these isolates. Majority of these patients recovered from the infection (65%) after proper antibiotic therapy.The ISApl1 transposable elements were not detected in these isolates. These isolates were found clonally unrelated when analyze by pulsed-field gel electrophoresis. The conjugation attempt to transfer mcr-1 to recipient’s E. coli J53 failed, Southern hybridization showed that mcr-1 was found located on chromosome in multiple copies. Conclusion This is the first case of mcr-1 in a human clinical isolate in Acinetobacter Baumnnii from India. These findings highlight the vertical transferability of colistin resistance by mcr-1 gene in Acinetobacter Baumnnii with the association of known some unknown insertion sequence located on chromosome. Strategies required to contain their spread and evolution of such genes. Disclosures All authors: No reported disclosures.

Construction of a genomic map ofMoraxella (Branhamella) catarrhalisATCC 25238 and physical mapping of virulence-associated genes

1999 ◽  
Vol 45 (4) ◽  
pp. 299-303 ◽  
Author(s):  
K T Nguyen ◽  
E J Hansen ◽  
M A Farinha
Keyword(s):  
Gel Electrophoresis ◽  
Moraxella Catarrhalis ◽  
Southern Hybridization ◽  
Physical Map ◽  
Outer Membrane Proteins ◽  
Restriction Enzymes ◽  
Pulsed Field Gel Electrophoresis ◽  
Respiratory Pathogens ◽  
Circular Chromosome ◽  
Pulsed Field

A physical genome map of the Moraxella catarrhalis type strain (ATCC 25238) has been constructed using pulsed field gel electrophoresis. Macrorestriction analyses of the genome of M. catarrhalis were performed by digestion with the restriction enzymes SmaI, NotI, and RsrII, which cleave the single circular chromosome into 9, 10, and 6 fragments, respectively. The chromosomal fragments generated by pulsed field gel electrophoresis were converted to a linkage map utilizing a combination of partial digestions, and cross-hybridizations. Moraxella catarrhalis, like a number of other respiratory pathogens, has a relatively small genome estimated at 1750 kilobase pairs or about 40% of the size of the Escherichia coli genome. The locations of the four ribosomal RNA operons (rrnLS) were determined by Southern hybridization and by digestion with I-CeuI endonuclease. A number of genes involved in virulence have been placed onto the physical map by Southern hybridization including those encoding the predominant outer-membrane proteins and the chromosomal gene encoding beta-lactamase.Key words: Moraxella catarrhalis, physical map, genome analysis, pulsed-field gel electrophoresis, virulence.

Physical and genetic map of the Spiroplasma kunkelii CR2-3x chromosome

2006 ◽  
Vol 52 (9) ◽  
pp. 857-867 ◽  
Author(s):  
Ellen L Dally ◽  
Thereza S.L Barros ◽  
Yan Zhao ◽  
ShaoPing Lin ◽  
Bruce A Roe ◽  
...  
Keyword(s):  
Genetic Map ◽  
Gel Electrophoresis ◽  
Southern Hybridization ◽  
Pulsed Field Gel Electrophoresis ◽  
Enzyme Analysis ◽  
Two Dimensional ◽  
Pulsed Field ◽  
Spiroplasma Kunkelii ◽  
Corn Stunt ◽  
Physical And Genetic Map

Spiroplasma kunkelii (class Mollicutes) is the characteristically helical, wall-less bacterium that causes corn stunt disease. A combination of restriction enzyme analysis, pulsed-field gel electrophoresis (PFGE), and Southern hybridization analysis was used to construct a physical and genetic map of the S. kunkelii CR2-3x chromosome. The order of restriction fragments on the map was determined by analyses of reciprocal endonuclease double digests employing I-CeuI, AscI, ApaI, EagI, SmaI, BssHII, BglI, and SalI; adjacent fragments were identified on two-dimensional pulsed-field electrophoresis gels. The size of the chromosome was estimated at 1550 kb. Oligonucleotide pairs were designed to prime the amplification of 26 S. kunkelii gene sequences in the polymerase chain reaction (PCR). Using PCR amplicons as probes, the locations of 27 S. kunkelii putative single-copy genes were positioned on the map by Southern hybridization analyses of chromosomal fragments separated in PFGE. The nucleotide sequence of the single ribosomal RNA operon was determined and its location mapped to a chromosomal segment bearing recognition sites for SalI, SmaI, EagI, and I-CeuI.Key words: Spiroplasma kunkelii CR2-3x, corn stunt spiroplasma, mollicutes, genome mapping, two-dimensional pulsed-field gel electrophoresis.

scholarly journals Plasmid-Borne Type E Neurotoxin Gene Clusters in Clostridium botulinum Strains

2013 ◽  
Vol 79 (12) ◽  
pp. 3856-3859 ◽  
Author(s):  
Zhen Zhang ◽  
Hannamari Hintsa ◽  
Ying Chen ◽  
Hannu Korkeala ◽  
Miia Lindström
Keyword(s):  
Gene Cluster ◽  
Gel Electrophoresis ◽  
Clostridium Botulinum ◽  
Southern Hybridization ◽  
Gene Clusters ◽  
Pulsed Field Gel Electrophoresis ◽  
Content Type ◽  
Pulsed Field ◽  
Large Plasmids

ABSTRACTA collection of 36Clostridium botulinumtype E strains was examined by pulsed-field gel electrophoresis (PFGE) and Southern hybridization with probes targeted tobotEandorfX1in the neurotoxin gene cluster. Three strains were found to contain neurotoxin subtype E1 gene clusters in large plasmids of about 146 kb in size.

scholarly journals Metallo-β-Lactamases in ClinicalPseudomonas Isolates in Taiwan and Identification of VIM-3, a Novel Variant of the VIM-2 Enzyme

2001 ◽  
Vol 45 (8) ◽  
pp. 2224-2228 ◽  
Author(s):  
Jing-Jou Yan ◽  
Po-Ren Hsueh ◽  
Wen-Chien Ko ◽  
Kwen-Tay Luh ◽  
Shu-Huei Tsai ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Amino Acid ◽  
Sequence Analysis ◽  
Gel Electrophoresis ◽  
Southern Hybridization ◽  
Pulsed Field Gel Electrophoresis ◽  
Pulsed Field ◽  
Novel Variant ◽  
Pcr Assays ◽  
Positive Results

ABSTRACT A total of 209 clinical isolates of Pseudomonas (193Pseudomonas aeruginosa, 10 P. putida, 4P. stutzeri, and 2 P. fluorescensisolates) with reduced susceptibilities to imipenem and/or ceftazidime were subjected to PCR assays with primers specific forbla IMP-1, bla IMP-2,bla VIM-1, and bla VIM-2and sequence analysis to identify the metallo-β-lactamases (MBLs) prevalent among these organisms in Taiwan; and 21 isolates gave positive results. Five isolates including two P. putida and three P. stutzeri isolates were found to carrybla IMP-1, and six isolates including fiveP. putida and one P. stutzeri isolates harboredbla VIM-2. The remaining 10 isolates wereP. aeruginosa, and all were found to carry a novel variant of bla VIM-2, designatedbla VIM-3. There are only two nucleotide differences between bla VIM-2 andbla VIM-3, leading to two amino acid alterations. Our findings indicate that VIM-2 and its variant have become the most prevalent metalloenzymes in Pseudomonas in Taiwan. Southern hybridization with thebla VIM-2-, bla VIM-3-, and bla IMP-1 -specific probes revealed that only two VIM-2-producing P. putida isolates appeared to carry the MBL gene on plasmids. Pulsed-field gel electrophoresis showed that six VIM-3-producing P. aeruginosa isolates and two IMP-1-producing P. stutzeri isolates were genetically related, suggesting that the spread of these MBL genes in Taiwan could be due to clonal dissemination as well as genetic exchange between different clones.

scholarly journals Genomic Complexity among Strains of the Facultative Photoheterotrophic Bacterium Rhodobacter sphaeroides

1999 ◽  
Vol 181 (5) ◽  
pp. 1684-1688 ◽  
Author(s):  
Kirsten Siedenburg Nereng ◽  
Samuel Kaplan
Keyword(s):  
Rhodobacter Sphaeroides ◽  
Gel Electrophoresis ◽  
Southern Hybridization ◽  
Pulsed Field Gel Electrophoresis ◽  
Restriction Endonucleases ◽  
Pulsed Field ◽  
Genomic Complexity ◽  
Genome Complexity

ABSTRACT Pulsed-field gel electrophoresis following the use of rare cutting restriction endonucleases together with Southern hybridization, using markers distributed on chromosomes I and II of Rhodobacter sphaeroides 2.4.1, has been used to examine approximately 25 strains of R. sphaeroides in an effort to assess the occurrence of genome complexity in these strains. The results suggest that genome complexity is widespread and is accompanied by substantial genomic heterogeneity.

scholarly journals Identification of 16S rRNA Methylase-Producing Acinetobacter baumannii Clinical Strains in North America

2007 ◽  
Vol 51 (11) ◽  
pp. 4209-4210 ◽  
Author(s):  
Yohei Doi ◽  
Jennifer M. Adams ◽  
Kunikazu Yamane ◽  
David L. Paterson
Keyword(s):  
North America ◽  
16S Rrna ◽  
Acinetobacter Baumannii ◽  
Gel Electrophoresis ◽  
Medical Center ◽  
Pulsed Field Gel Electrophoresis ◽  
Aminoglycoside Resistance ◽  
Pulsed Field ◽  
Clinical Strains ◽  
16S Rrna Methylase

ABSTRACT Five highly amikacin-resistant Acinetobacter baumannii isolates were collected at a medical center in Pennsylvania. The aminoglycoside resistance was due to the production of the 16S rRNA methylase ArmA. Two of the isolates coproduced OXA-23 β-lactamase and were highly resistant to carbapenems as well. The isolates were genetically closely related by pulsed-field gel electrophoresis.

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