scholarly journals Molecular Characterization of a Cephamycin-Hydrolyzing and Inhibitor-Resistant Class A β-Lactamase, GES-4, Possessing a Single G170S Substitution in the Ω-Loop

2004 ◽  
Vol 48 (8) ◽  
pp. 2905-2910 ◽  
Author(s):  
Jun-ichi Wachino ◽  
Yohei Doi ◽  
Kunikazu Yamane ◽  
Naohiro Shibata ◽  
Tetsuya Yagi ◽  
...  
Keyword(s):  
Neonatal Intensive Care ◽  
Gene Cassette ◽  
Single Amino Acid ◽  
Sequencing Analysis ◽  
Loop Region ◽  
Class 1 ◽  
Different Strains ◽  
Hydrolysis Of ◽  
Cloning Experiment

ABSTRACT The nosocomial spread of six genetically related Klebsiella pneumoniae strains producing GES-type β-lactamases was found in a neonatal intensive care unit, and we previously reported that one of the six strains, strain KG525, produced a new β-lactamase, GES-3. In the present study, the molecular mechanism of cephamycin resistance observed in strain KG502, one of the six strains described above, was investigated. This strain was found to produce a variant of GES-3, namely, GES-4, which was responsible for resistance to both cephamycins (cefoxitin MIC, >128 μg/ml) and β-lactamase inhibitors (50% inhibitory concentration of clavulanic acid, 15.2 ± 1.7 μM). The GES-4 enzyme had a single G170S substitution in the Ω-loop region compared with the GES-3 sequence. This single amino acid substitution was closely involved with the augmented hydrolysis of cephamycins and carbapenems and the decreased affinities of β-lactamase inhibitors to GES-4. A cloning experiment and sequencing analysis revealed that strain KG502 possesses duplicate bla GES-4 genes mediated by two distinct class 1 integrons with similar gene cassette configurations. Moreover, the genetic environments of the bla GES-4 genes found in strain KG502 were almost identical to that of bla GES-3 in strain KG525. From these findings, these two phenotypically different strains were suggested to belong to a clonal lineage. The bla GES-4 gene found in strain KG502 might well emerge from a point mutation in the bla GES-3 gene harbored by its ancestor strains, such as strain KG525, under heavy antibiotic stress in order to acquire extended properties of resistance to cephamycins and carbapenems.

scholarly journals Characterization of a Novel Plasmid-Mediated Cephalosporinase (CMY-9) and Its Genetic Environment in an Escherichia coli Clinical Isolate

2002 ◽  
Vol 46 (8) ◽  
pp. 2427-2434 ◽  
Author(s):  
Yohei Doi ◽  
Naohiro Shibata ◽  
Keigo Shibayama ◽  
Kazunari Kamachi ◽  
Hiroshi Kurokawa ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Gene Cassette ◽  
Single Amino Acid ◽  
Common Region ◽  
Class 1 Integron ◽  
Class 1 ◽  
The Common ◽  
High Level

ABSTRACT An Escherichia coli strain, HKYM68, which showed resistance to broad-spectrum cephalosporins was isolated from a sputum specimen in Japan. The high-level resistance of the strain to ceftazidime, cefpirome, and moxalactam was carried by a self-transferable plasmid. The β-lactamase gene responsible for the resistance was cloned and sequenced. The deduced amino acid sequence of this gene product, CMY-9, had a single amino acid substitution (E85D), the residue reported to be part of the recognition site for the R1 side chain of β-lactams, compared with the amino acid sequence of CMY-8 and also had 78% identity with the amino acid sequence of CepH, a chromosomal cephalosporinase of Aeromonas hydrophila. A sul1-type class 1 integron containing an aacA1-orfG gene cassette was identified upstream of bla CMY-9 and ended with a truncated 3′ conserved segment. The following 2.1 kb was almost identical to the common region of integrons In6 and In7 and the integron of pSAL-1, except that orf513 encoding a putative transposase was identified instead of orf341 due to addition of a single nucleotide. bla CMY-9 was closely located downstream of the end of the common region. These observations are indicative of the exogenous derivation of bla CMY-9 from some environmental microorganisms such as aeromonads.

scholarly journals Genetic Context and Biochemical Characterization of the IMP-18 Metallo-β-Lactamase Identified in aPseudomonas aeruginosaIsolate from the United States

2010 ◽  
Vol 55 (1) ◽  
pp. 140-145 ◽  
Author(s):  
Luisa Borgianni ◽  
Silvia Prandi ◽  
Laurie Salden ◽  
Gisela Santella ◽  
Nancy D. Hanson ◽  
...  
Keyword(s):  
United States ◽  
Biochemical Characterization ◽  
Gene Cassette ◽  
The United States ◽  
Turnover Rates ◽  
Reading Frame ◽  
Class 1 ◽  
Original Array ◽  
Genetic Context

ABSTRACTThe production of metallo-β-lactamase (MBL) is an important mechanism of resistance to β-lactam antibiotics, including carbapenems. Despite the discovery and emergence of many acquired metallo-β-lactamases, IMP-type determinants (now counting at least 27 variants) remain the most prevalent in some geographical areas. In Asian countries, and notably Japan, IMP-1 and its closely related variants are most widespread. Some other variants have been detected in other countries and show either an endemic (e.g., IMP-13 in Italy) or sporadic (e.g., IMP-12 in Italy or IMP-18 in the United States) occurrence. The IMP-18-producingPseudomonas aeruginosastrain PS 297 from the southwestern United States carried at least two class 1 integrons. One was identical to In51, while the other, named In133and carrying theblaIMP-18gene cassette in the third position, showed an original array of five gene cassettes, includingaacA7,qacF,aadA1, and an unknown open reading frame (ORF). Interestingly. In133differed significantly from In96, theblaIMP-18-carrying integron identified in aP. aeruginosaisolate from Mexico. The meropenem and ertapenem MIC values were much lower forEscherichia colistrains producing IMP-18 (0.06 and 0.12 μg/ml, respectively) than for strains producing IMP-1 (2 μg/ml for each). Kinetic data obtained with the purified enzyme revealed lower turnover rates of IMP-18 than of other IMP-type enzymes with most substrates.

scholarly journals Characterization of In53, a Class 1 Plasmid- and Composite Transposon-Located Integron of Escherichia coli Which Carries an Unusual Array of Gene Cassettes

2001 ◽  
Vol 183 (1) ◽  
pp. 235-249 ◽  
Author(s):  
Thierry Naas ◽  
Yuzuru Mikami ◽  
Tamae Imai ◽  
Laurent Poirel ◽  
Patrice Nordmann
Keyword(s):  
Escherichia Coli ◽  
Resistance Gene ◽  
Gene Cassette ◽  
Promoter Sequence ◽  
Class 1 Integron ◽  
Integrase Gene ◽  
Chloramphenicol Resistance ◽  
Gene Cassettes ◽  
Class 1

ABSTRACT Further characterization of the genetic environment of the gene encoding the Escherichia coli extended-spectrum β-lactamase, bla VEB-1, revealed the presence of a plasmid-located class 1 integron, In53, which carried eight functional resistance gene cassettes in addition tobla VEB-1. While the aadB and the arr-2 gene cassettes were identical to those previously described, the remaining cassettes were novel: (i) a novel nonenzymatic chloramphenicol resistance gene of the cmlAfamily, (ii) a qac allele encoding a member of the small multidrug resistance family of proteins, (iii) a cassette,aacA1b/orfG, which encodes a novel 6′-N-acetyltransferase, and (iv) a fused gene cassette,oxa10/aadA1, which is made of two cassettes previously described as single cassettes. In addition, oxa10 andaadA1 genes were expressed from their own promoter sequence present upstream of the oxa10 cassette.arr-2 coded for a protein that shared 54% amino acid identity with the rifampin ADP-ribosylating transferase encoded by thearr-1 gene from Mycobacterium smegmatisDSM43756. While in M. smegmatis, the main inactivated compound was 23-ribosyl-rifampin, the inactivated antibiotic recovered from E. coli culture was 23-O-ADP-ribosyl-rifampin. The integrase gene of In53 was interrupted by an IS26 insertion sequence, which was also present in the 3′ conserved segment. Thus, In53 is a truncated integron located on a composite transposon, named Tn2000, bounded by two IS26 elements in opposite orientations. Target site duplication at both ends of the transposon indicated that the integron likely was inserted into the plasmid through a transpositional process. This is the first description of an integron located on a composite transposon.

scholarly journals Phenotypic and Enzymatic Comparative Analysis of the Novel KPC Variant KPC-5 and Its Evolutionary Variants, KPC-2 and KPC-4

2008 ◽  
Vol 53 (2) ◽  
pp. 557-562 ◽  
Author(s):  
Daniel J. Wolter ◽  
Philip M. Kurpiel ◽  
Neil Woodford ◽  
Marie-France I. Palepou ◽  
Richard V. Goering ◽  
...  
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequences ◽  
Single Amino Acid ◽  
Upstream Region ◽  
The Novel ◽  
Carbapenem Resistant ◽  
Flanking Regions ◽  
Hydrolysis Of ◽  
Additional Amino Acid

ABSTRACT A novel Klebsiella pneumoniae carbapenemase (KPC) variant, designated bla KPC-5, was discovered in a carbapenem-resistant Pseudomonas aeruginosa clinical isolate from Puerto Rico. Characterization of the upstream region of bla KPC-5 showed significant differences from the flanking regions of other bla KPC variants. Comparison of amino acid sequences with those of other KPC enzymes revealed that KPC-5 was an intermediate between KPC-2 and KPC-4, differing from KPC-2 by a single amino acid substitution (Pro103→Arg), while KPC-4 contained Pro103→Arg plus an additional amino acid change (Val239→Gly). Transformation studies with an Escherichia coli recipient strain showed differences in the properties of the KPC variants. KPC-4 and KPC-5 both had pIs of 7.65, in contrast with the pI of 6.7 for KPC-2. KPC-2 transformants were less susceptible to the carbapenems than KPC-4 and KPC-5 transformants. These data correlated with higher rates of imipenem hydrolysis for KPC-2 than for KPC-4 and KPC-5. However, KPC-4 and KPC-5 transformants had higher ceftazidime MICs, and the enzymes from these transformants had slightly better hydrolysis of this drug than KPC-2. KPC-4 and KPC-5 were more sensitive than KPC-2 to inhibition by clavulanic acid in both susceptibility testing and hydrolysis assays. Thus, KPC enzymes may be evolving through stepwise mutations to alter their spectra of activity.

Characterization of Gating and Peptide Block of mSlo, a Cloned Calcium-Dependent Potassium Channel

1997 ◽  
Vol 78 (6) ◽  
pp. 2937-2950 ◽  
Author(s):  
Deirdre A. Sullivan ◽  
Mats H. Holmqvist ◽  
Irwin B. Levitan
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Potassium Channel ◽  
Critical Role ◽  
Single Amino Acid ◽  
Significant Shift ◽  
Triple Mutant ◽  
Calcium Dependent ◽  
Loop Region

Sullivan, Deirdre A., Mats H. Holmqvist, and Irwin B. Levitan. Characterization of gating and peptide block of mSlo, a cloned calcium-dependent potassium channel. J. Neurophysiol. 78: 2937–2950, 1997. The 20 amino acid Shaker inactivation peptide blocks mSlo, a cloned calcium-dependent potassium channel. Changing the charge and degree of hydrophobicity of the peptide alters its blocking kinetics. A “triple mutant” mSlo channel was constructed in which three amino acids (T256, S259, and L262), equivalent to those identified as part of the peptide's receptor site in the S4–S5 cytoplasmic loop region of the Shaker channel, were mutated simultaneously to alanines. These mutations produce only limited changes in the channel's susceptibility to block by a series of peptides of varying charge and hydrophobicity but do alter channel gating. The triple mutant channel shows a significant shift in its calcium-activation curve as compared with the wild-type channel. Analysis of the corresponding single amino acid mutations shows that mutation at position L262 causes the most dramatic change in mSlo gating. These results suggest that the three amino acids mutated in the mSlo S4–S5 loop may contribute to, but are not essential for, peptide binding. On the other hand, they do play a critical role in the channel's calcium-sensing mechanism.

scholarly journals Molecular Characterization of Class 3 Integrons from Delftia spp.

2007 ◽  
Vol 189 (17) ◽  
pp. 6276-6283 ◽  
Author(s):  
Hai Xu ◽  
Julian Davies ◽  
Vivian Miao
Keyword(s):  
Antibiotic Resistance ◽  
Antibiotic Resistance Genes ◽  
Gene Cassette ◽  
Integrase Gene ◽  
Delftia Acidovorans ◽  
Sequence Identity ◽  
Chromosomal Genes ◽  
Class 1 ◽  
Delftia Tsuruhatensis

ABSTRACT Two environmental strains, Delftia acidovorans C17 and Delftia tsuruhatensis A90, were found to carry class 3 integrons, which have seldom been reported and then only from pathogens in which they are associated with antibiotic resistance genes. The Delftia integrons comprised a highly conserved class 3 integrase gene, upstream and oppositely oriented from a set of three or four gene cassettes that encoded unidentified functions. The A90 integron had one more gene cassette than the C17 integron, but the two were otherwise the same; furthermore, they were located within regions of sequence identity in both strains and linked to chromosomal genes. A screen of other Delftia and related strains did not reveal the presence of additional class 3 integrons. The observations suggest that these integrons were horizontally transferred to Delftia as part of a larger region and reside as chromosomal elements that probably predate transposon dissemination, as has been proposed for certain class 1 integrons.

INTEGRON DIVERSITY IN BLAVIM-2-CARRYING CARBAPENEM-RESISTANT CLINICAL PSEUDOMONAS AERUGINOSA ISOLATES

2019 ◽  
Vol 64 (8) ◽  
pp. 497-502 ◽  
Author(s):  
T. A. Savinova ◽  
Yu. A. Bocharova ◽  
A. V. Lazareva ◽  
I. V. Chebotar ◽  
N. A. Mayanskiy
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Gene Cassette ◽  
Variable Regions ◽  
Class 1 Integrons ◽  
Carbapenem Resistant ◽  
Class 1 ◽  
Encoding Gene ◽  
Clonal Spread ◽  
Pathogen Populations

The growing prevalence of metallo-β-lactamase (MBL)-producing Pseudomonas aeruginosa in nosocomial pathogen populations has been attributed to their clonal spread, and/or horizontal transfer of MBL determinants in mobile genetic elements, including integrons. To characterize the genetic background of the beta-lactamase VIM-2 encoding gene in the population of carbapenem-resistant (Carba-R) P. aeruginosa clinical isolates.The detection of class 1 integrons was performed by PCR. Typing of the class 1 integrons containing the blaVIM gene cassette was performed by the PCR-restriction fragment length polymorphism (RFLP) approach followed by sequencing of variable regions of class 1 integrons. Five types of the blaVIM-2-carrying integrons were identified: ST654-isolates accounting for more than 50% of the Carba-R population harbored In56; ST235-isolates contained In559 (26% Carba-R isolates); ST111-isolates (19% Carba-R isolates) were characterized by carrying In59-like integron; two ST235-isolates harbored In59 and In249 each. Except In56, carrying the only blaVIM-2-gene cassette, all other identified integron types harbored the genes of resistance to trimethoprim and/or aminoglycosides. No new types of integrons were identified in the P. aeruginosa clinical isolates. The observed correlation of the integron type with specific STs indicates a clonal dissemination of significant resistance determinant producers - ST111, ST654 and ST235 epidemic lines. The features of the integron variable regions can be used for the epidemiological characterization of clinical P. aeruginosa isolates.

scholarly journals Genetic characterization of multidrug resistance in Shigella spp. from Japan

2006 ◽  
Vol 55 (12) ◽  
pp. 1685-1691 ◽  
Author(s):  
Ashraf M. Ahmed ◽  
Kimi Furuta ◽  
Kei Shimomura ◽  
Yoshio Kasama ◽  
Tadashi Shimamoto
Keyword(s):  
Shigella Flexneri ◽  
Blot Hybridization ◽  
Southern Blot Hybridization ◽  
Shigella Sonnei ◽  
Sequencing Analysis ◽  
Class 1 Integron ◽  
Gene Cassettes ◽  
Class 1 ◽  
Shigella Spp

This study characterized the genetic basis of antimicrobial resistance of a number of Shigella spp. isolated from humans from 2000 to 2004 in Hiroshima prefecture, Japan. A total of 26 isolates of Shigella spp. were included in this study. Antimicrobial susceptibility tests revealed high levels of resistance, especially to ampicillin, streptomycin, trimethoprim, tetracycline, nalidixic acid and ciprofloxacin. PCR and DNA sequencing were used for screening and characterization of antibiotic-resistance determinants. PCR sequencing analysis revealed the presence of only one type of class 1 integron in one isolate of Shigella sonnei. This class 1 integron was 1904 bp and contained two gene cassettes: a probable esterase/lipase (estX) and aadA1, which confers resistance to streptomycin and spectinomycin. Two types of class 2 integron were identified in this study. One was the classic type (2158 bp) and carried the three conserved resistance gene cassettes of the class 2 integron, dfrA1, sat1 and aadA1, which confer resistance to trimethoprim, streptothricin and streptomycin/spectinomycin, respectively. This type was detected in both Shigella sonnei (14 isolates) and Shigella flexneri (five isolates). The other type was shorter (1313 bp) and carried only two gene cassettes, dfrA1 and sat1. This integron was detected in a single isolate of Shigella sonnei. PFGE patterns showed limited diversity within clusters of the same species. Furthermore, an extended-spectrum β-lactamase gene, bla OXA-30, which confers resistance to ampicillin, was characterized in all isolates of Shigella flexneri except the oldest strain, which was isolated in 2000. Southern blot hybridization and conjugation experiments showed that bla OXA-30 was located in the chromosome.

scholarly journals Molecular and epidemiologic characterization of the diphtheria outbreak in Venezuela

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Ricardo A. Strauss ◽  
Laura Herrera-Leon ◽  
Ana C. Guillén ◽  
Julio S. Castro ◽  
Eva Lorenz ◽  
...  
Keyword(s):  
Core Genome ◽  
Spanning Trees ◽  
Case Fatality ◽  
Minimum Spanning Trees ◽  
Sequencing Analysis ◽  
Age Group ◽  
Coverage Problems ◽  
Different Strains ◽  
Clinical Records

AbstractIn 2016, Venezuela faced a large diphtheria outbreak that extended until 2019. Nasopharyngeal or oropharyngeal samples were prospectively collected from 51 suspected cases and retrospective data from 348 clinical records was retrieved from 14 hospitals between November 2017 and November 2018. Confirmed pathogenic Corynebactrium isolates were biotyped. Multilocus Sequence Typing (MLST) was performed followed by next-generation-based core genome-MLST and minimum spanning trees were generated. Subjects between 10 and 19 years of age were mostly affected (n = 95; 27.3%). Case fatality rates (CFR) were higher in males (19.4%), as compared to females (15.8%). The highest CFR (31.1%) was observed among those under 5, followed by the 40 to 49 age-group (25.0%). Nine samples corresponded to C. diphtheriae and 1 to C. ulcerans. Two Sequencing Types (ST), ST174 and ST697 (the latter not previously described) were identified among the eight C. diphtheriae isolates from Carabobo state. Cg-MLST revealed only one cluster also from Carabobo. The Whole Genome Sequencing analysis revealed that the outbreak seemed to be caused by different strains with C. diphtheriae and C. ulcerans coexisting. The reemergence and length of this outbreak suggest vaccination coverage problems and an inadequate control strategy.

Sign in / Sign up

Export Citation Format

Share Document