( 2003 )
Cloning and sequencing of the histidine decarboxylase genes of gram-negative, histamine-producing bacteria and their application in detection and identification of these organisms in fish.
PMID : 12732523 : DOI : 10.1128/aem.69.5.2568-2579.2003 PMC : PMC154508
The use of molecular tools for early and rapid detection of gram-negative histamine-producing bacteria is important for preventing the accumulation of histamine in fish products. To date, no molecular detection or identification system for gram-negative histamine-producing bacteria has been developed. A molecular method that allows the rapid detection of gram-negative histamine producers by PCR and simultaneous differentiation by single-strand conformation polymorphism (SSCP) analysis using the amplification product of the histidine decarboxylase genes (hdc) was developed. A collection of 37 strains of histamine-producing bacteria (8 reference strains from culture collections and 29 isolates from fish) and 470 strains of non-histamine-producing bacteria isolated from fish were tested. Histamine production of bacteria was determined by paper chromatography and confirmed by high-performance liquid chromatography. Among 37 strains of histamine-producing bacteria, all histidine-decarboxylating gram-negative bacteria produced a PCR product, except for a strain of Citrobacter braakii. In contrast, none of the non-histamine-producing strains (470 strains) produced an amplification product. Specificity of the amplification was further confirmed by sequencing the 0.7-kbp amplification product. A phylogenetic tree of the isolates constructed using newly determined sequences of partial hdc was similar to the phylogenetic tree generated from 16S ribosomal DNA sequences. Histamine accumulation occurred when PCR amplification of hdc was positive in all of fish samples tested and the presence of powerful histamine producers was confirmed by subsequent SSCP identification. The potential application of the PCR-SSCP method as a rapid monitoring tool is discussed.
( 2003 )
Mercury resistance determinants related to Tn21, Tn1696, and Tn5053 in enterobacteria from the preantibiotic era.
PMID : 12604550 : DOI : 10.1128/aac.47.3.1115-1119.2003 PMC : PMC149298
Three mer transposons from the Murray collection of preantibiotic enterobacteria show >99% sequence identity to current isolates. Tn5073 is most closely related to Tn5036 and Tn1696, and Tn5074 is most closely related to Tn5053. Tn5075 is most closely related to Tn21 but lacks integron In2 and is flanked by insertion elements.
( 2002 )
Evolution of the gyrB gene and the molecular phylogeny of Enterobacteriaceae: a model molecule for molecular systematic studies.
PMID : 11931166 : DOI : 10.1099/00207713-52-2-531
Phylogenetic trees showing the evolutionary relatedness of Enterobacteriaceae based upon gyrB and 16S rRNA genes were compared. Congruence among trees of these molecules indicates that the genomes of these species are not completely mosaic and that molecular systematic studies can be carried out. Phylogenetic trees based on gyrB sequences appeared to be more reliable at determining relationships among Serratia species than trees based on 16S rRNA gene sequences. gyrB sequences from Serratia species formed a monophyletic group validated by significant bootstrap values. Serratia fonticola had the most deeply branching gyrB sequence in the Serratia monophyletic group, which was consistent with its atypical phenotypic characteristics. Klebsiella and Enterobacter genera seemed to be polyphyletic, but the branching patterns of gyrB and 16S rRNA gene trees were not congruent. Enterobacter aerogenes was grouped with Klebsiella pneumoniae on the gyrB phylogenetic tree, which supports that this species could be transferred to the Klebsiella genus. Unfortunately, 16S rRNA and gyrB phylogenetic trees gave conflicting evolutionary relationships for Citrobacter freundii because of its unusual gyrB evolutionary process. gyrB lateral gene transfer was suspected for Hafnia alvei. Saturation of gyrB genes was observed by the pairwise comparison of Proteus spp., Providencia alcalifaciens and Morganella morganii sequences. Depending on their level of variability, 16S rRNA gene sequences were useful for describing phylogenetic relationships between distantly related Enterobacteriaceae, whereas gyrB sequence comparison was useful for inferring intra- and some intergeneric relationships.
( 2000 )
Phosphorylation of nucleosides by the mutated acid phosphatase from Morganella morganii.
PMID : 10877772 : DOI : 10.1128/aem.66.7.2811-2816.2000 PMC : PMC92077
A novel nucleoside phosphorylation process using the food additive pyrophosphate as the phosphate source was investigated. The Morganella morganii gene encoding a selective nucleoside pyrophosphate phosphotransferase was cloned. It was identical to the M. morganii PhoC acid phosphatase gene. Sequential in vitro random mutagenesis was performed on the gene by error-prone PCR to construct a mutant library. The mutant library was introduced into Escherichia coli, and the transformants were screened for the production of 5'-IMP. One mutated acid phosphatase with an increased phosphotransferase reaction yield was obtained. With E. coli overproducing the mutated acid phosphatase, 101 g of 5'-IMP per liter (192 mM) was synthesized from inosine in an 88% molar yield. This improvement was achieved with two mutations, Gly to Asp at position 92 and Ile to Thr at position 171. A decreased K(m) value for inosine was responsible for the increased productivity.
de Massis MR,
( 2000 )
TEM-72, a new extended-spectrum beta-lactamase detected in Proteus mirabilis and Morganella morganii in Italy.
PMID : 10952610 : DOI : 10.1128/aac.44.9.2537-2539.2000 PMC : PMC90100
A new natural TEM-2 derivative, named TEM-72, was identified in a Proteus mirabilis strain and in a Morganella morganii strain isolated in Italy in 1999. Compared to TEM-1, TEM-72 contains the following amino acid substitutions: Q39K, M182T, G238S, and E240K. Kinetic analysis showed that TEM-72 exhibits an extended-spectrum activity, including activity against oxyimino-cephalosporins and aztreonam. Expression of bla(TEM-72) in Escherichia coli was capable of decreasing the host susceptibility to the above drugs.
( 1999 )
Phylogenetic analysis of L4-mediated autogenous control of the S10 ribosomal protein operon.
PMID : 10498727 : PMC : PMC103642
We investigated the regulation of the S10 ribosomal protein (r-protein) operon among members of the gamma subdivision of the proteobacteria, which includes Escherichia coli. In E. coli, this 11-gene operon is autogenously controlled by r-protein L4. This regulation requires specific determinants within the untranslated leader of the mRNA. Secondary structure analysis of the S10 leaders of five enterobacteria (Salmonella typhimurium, Citrobacter freundii, Yersinia enterocolitica, Serratia marcescens, and Morganella morganii) and two nonenteric members of the gamma subdivision (Haemophilus influenzae and Vibrio cholerae) shows that these foreign leaders share significant structural homology with the E. coli leader, particularly in the region which is critical for L4-mediated autogenous control in E. coli. Moreover, these heterologous leaders produce a regulatory response to L4 oversynthesis in E. coli. Our results suggest that an E. coli-like L4-mediated regulatory mechanism may operate in all of these species. However, the mechanism is not universally conserved among the gamma subdivision members, since at least one, Pseudomonas aeruginosa, does not contain the required S10 leader features, and its leader cannot provide the signals for regulation by L4 in E. coli. We speculate that L4-mediated autogenous control developed during the evolution of the gamma branch of proteobacteria.
( 1999 )
Cloning, sequence analyses, expression, and distribution of ampC-ampR from Morganella morganii clinical isolates.
PMID : 10103179 : PMC : PMC89205
Shotgun cloning experiments with restriction enzyme-digested genomic DNA from Morganella morganii 1, which expresses high levels of cephalosporinase, into the pBKCMV cloning vector gave a recombinant plasmid, pPON-1, which encoded four entire genes: ampC, ampR, an hybF family gene, and orf-1 of unknown function. The deduced AmpC beta-lactamase of pI 7.6 shared structural and functional homologies with AmpC from Citrobacter freundii, Escherichia coli, Yersinia enterocolitica, Enterobacter cloacae, and Serratia marcescens. The overlapping promoter organization of ampC and ampR, although much shorter in M. morganii than in the other enterobacterial species, suggested similar AmpR regulatory properties. The MICs of beta-lactams for E. coli MC4100 (ampC mutant) harboring recombinant plasmid pACYC184 containing either ampC and ampR (pAC-1) or ampC (pAC-2) and induction experiments showed that the ampC gene of M. morganii 1 was repressed in the presence of ampR and was activated when a beta-lactam inducer was added. Moreover, transformation of M. morganii 1 or of E. coli JRG582 (delta ampDE) harboring ampC and ampR with a recombinant plasmid containing ampD from E. cloacae resulted in a decrease in the beta-lactam MICs and an inducible phenotype for M. morganii 1, thus underlining the role of an AmpD-like protein in the regulation of the M. morganii cephalosporinase. Fifteen other M. morganii clinical isolates with phenotypes of either low-level inducible cephalosporinase expression or high-level constitutive cephalosporinase expression harbored the same ampC-ampR organization, with the hybF and orf-1 genes surrounding them; the organization of these genes thus differed from those of ampC-ampR genes in C. freundii and E. cloacae, which are located downstream from the fumarate operon. Finally, an identical AmpC beta-lactamase (DHA-1) was recently identified as being plasmid encoded in Salmonella enteritidis, and this is confirmatory evidence of a chromosomal origin of the plasmid-mediated cephalosporinases.
( 2007 )
Characterization of In3Mor, a new integron carrying VIM-1 metallo-beta-lactamase and sat1 gene, from Morganella morganii.
PMID : 17341470 : DOI : 10.1093/jac/dkm020
A carbapenem-resistant Morganella morganii clinical isolate that was phenotypically metallo-beta-lactamase (MBL)-positive was recovered from a Greek patient. The aim of the study was to analyse the structure of the integron containing the MBL gene. MICs were determined by the broth microdilution method. PCR assays and nucleotide sequencing were used for identification of bla gene types and mapping of the integron carrying the MBL gene. The location of the MBL allele was investigated by mating experiments and plasmid analysis as well as by Southern blotting of the plasmid extract and gene-specific hybridization with a bla(VIM-1) probe. The strain contained In3Mor, a novel class 1 integron carrying a carbapenemase gene (bla(VIM-1)) associated with a trimethoprim (dfrA1), a streptothricin (sat1) and two aminoglycoside resistance genes (aacA7 and aadA1). Conjugation experiments failed to detect a transferable MBL determinant and plasmid DNA was not visualized. The chromosomal location of bla(VIM-1) was confirmed after hybridization of the chromosomal band with the bla(VIM-1) probe. Production of a VIM-type MBL in a M. morganii clinical isolate is documented in this study for the first time. Also, the dfrA1-sat1-aadA1 array which is typically described in the variable region of class 2 integrons consistent with that on Tn7 transposons, is originally detected herein in a class 1 integron.
( 2007 )
Histidine decarboxylases and their role in accumulation of histamine in tuna and dried saury.
PMID : 17220267 : DOI : 10.1128/AEM.01907-06 PMC : PMC1828783
Histamine-producing bacteria (HPB) such as Photobacterium phosphoreum and Raoultella planticola possess histidine decarboxylase (HDC), which converts histidine into histamine. Histamine fish poisoning (HFP) is attributable to the ingestion of fish containing high levels of histamine produced by HPB. Because freezing greatly decreases the histamine-producing ability of HPB, especially of P. phosphoreum, it has been speculated that HFP is caused by HDC itself from HPB cells autolyzing during frozen storage, even when HPB survive frozen storage. Here we constructed recombinant HDCs of P. phosphoreum, Photobacterium damselae, R. planticola, and Morganella morganii and investigated the ability of HDCs to produce sufficient histamine to cause HFP. To elucidate the character of these HDCs, we examined the specific activity of each recombinant HDC at various temperatures, pH levels, and NaCl concentrations. Further, we also investigated the stability of each HDC under different conditions (in reaction buffer, tuna, and dried saury) at various temperatures. P. damselae HDC readily produced sufficient histamine to cause HFP in fish samples. We consider that if HDC is implicated as an independent cause of HFP in frozen-thawed fish, the most likely causative agent is HDC of P. damselae.
De Angelis AA,
( 2006 )
Structure determination of a membrane protein with two trans-membrane helices in aligned phospholipid bicelles by solid-state NMR spectroscopy.
PMID : 16967977 : DOI : 10.1021/ja063640w PMC : PMC3236029
The structure of the membrane protein MerFt was determined in magnetically aligned phospholipid bicelles by solid-state NMR spectroscopy. With two trans-membrane helices and a 10-residue inter-helical loop, this truncated construct of the mercury transport membrane protein MerF has sufficient structural complexity to demonstrate the feasibility of determining the structures of polytopic membrane proteins in their native phospholipid bilayer environment under physiological conditions. PISEMA, SAMMY, and other double-resonance experiments were applied to uniformly and selectively (15)N-labeled samples to resolve and assign the backbone amide resonances and to measure the associated (15)N chemical shift and (1)H-(15)N heteronuclear dipolar coupling frequencies as orientation constraints for structure calculations. (1)H/(13)C/(15)N triple-resonance experiments were applied to selectively (13)C'- and (15)N-labeled samples to complete the resonance assignments, especially for residues in the nonhelical regions of the protein. A single resonance is observed for each labeled site in one- and two-dimensional spectra. Therefore, each residue has a unique conformation, and all protein molecules in the sample have the same three-dimensional structure and are oriented identically in planar phospholipid bilayers. Combined with the absence of significant intensity near the isotropic resonance frequency, this demonstrates that the entire protein, including the loop and terminal regions, has a well-defined, stable structure in phospholipid bilayers.
( 2006 )
Integron presence in a multiresistant Morganella morganii isolate.
PMID : 16690260 : DOI : 10.1016/j.ijantimicag.2006.01.006
A multiresistant strain of Morganella morganii was isolated from a patient affected by several severe pathologies. The isolate was found to be resistant to the following antimicrobials: ampicillin, nalidixic acid, cefalothin, cefoxitin, ceftriaxone, ciprofloxacin, chloramphenicol, streptomycin, erythromycin, gentamicin, novobiocin, penicillin, rifampicin, tetracycline and violet crystal. Mechanisms leading to this multiresistance were studied. Porins of M. morganii multiresistant and wild-type strains were analysed by sodium dodecylsulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and were characterised by their ability to form channels in planar black lipid bilayers. The channels formed by porins from multiresistant and susceptible strains suggested that the porins of the multiresistant strain were not responsible for resistance. A 6.6 kb plasmid (pML2003) was detected, isolated and studied. pML2003 included two integrons. Direct sequencing revealed that one of the integrons contained two cassettes, aminoglycoside adenyltransferase (aadB) and chloramphenicol acetyltransferase (catB3) conferring resistance to aminoglycosides and chloramphenicol, respectively. The second integron contained carbenicillinase (blaP1b) and adenyltransferase (aadA2), which confer resistance to beta-lactamases and streptomycin, respectively.
( 2006 )
Morganella psychrotolerans sp. nov., a histamine-producing bacterium isolated from various seafoods.
PMID : 17012582 : DOI : 10.1099/ijs.0.64357-0
Mesophilic Morganella morganii (n=6) and psychrotolerant M. morganii-like isolates from various seafoods (n=13), as well as clinical M. morganii isolates (n=3), were characterized by using a polyphasic approach including multi-locus sequencing. Based on the phylogenetic analysis, the 22 strains were divided into two distinct groups comprising mesophilic and psychrotolerant isolates, respectively. This classification was supported by DNA-DNA hybridization studies, whereby a psychrotolerant isolate (strain U2/3(T)) showed 41.0 and 17.8 % relatedness to the type strains of the mesophilic species Morganella morganii subsp. morganii (strain LMG 7874(T)) and Morganella morganii subsp. sibonii (strain DSM 14850(T)), respectively. Analysis of the 16S rRNA gene sequences showed a similarity of 98.6 % between mesophilic and psychrotolerant isolates. However, fragments of seven protein-encoding housekeeping genes (atpD, dnaN, gyrB, hdc, infB, rpoB and tuf) all showed less than 90.9 % sequence similarity between the two groups. The psychrotolerant isolates grew at 0-2 degrees C and also differed from the mesophilic M. morganii isolates with respect to growth at 37 degrees C and in 8.5 % (w/v) NaCl and fermentation of d-galactose. The psychrotolerant strains appear to represent a novel species, for which the name Morganella psychrotolerans sp. nov. is proposed. The type strain is U2/3(T) (=LMG 23374(T)=DSM 17886(T)).
( 2005 )
NMR structure determination of a membrane protein with two transmembrane helices in micelles: MerF of the bacterial mercury detoxification system.
PMID : 15794657 : DOI : 10.1021/bi048095v
The three-dimensional backbone structure of a membrane protein with two transmembrane helices in micelles was determined using solution NMR methods that rely on the measurement of backbone (1)H-(15)N residual dipolar couplings (RDCs) from samples of two different constructs that align differently in stressed polyacrylamide gels. Dipolar wave fitting to the (1)H-(15)N RDCs determines the helical boundaries based on periodicity and was utilized in the generation of supplemental dihedral restraints for the helical segments. The (1)H-(15)N RDCs and supplemental dihedral restraints enable the determination of the structure of the helix-loop-helix core domain of the mercury transport membrane protein MerF with a backbone RMSD of 0.58 A. Moreover, the fold of this polypeptide demonstrates that the two vicinal pairs of cysteine residues, shown to be involved in the transport of Hg(II) across the membrane, are exposed to the cytoplasm. This finding differs from earlier structural and mechanistic models that were based primarily on the somewhat atypical hydropathy plot for MerF and related transport proteins.
( 2005 )
AcrAB efflux pump plays a role in decreased susceptibility to tigecycline in Morganella morganii.
PMID : 15673770 : DOI : 10.1128/AAC.49.2.791-793.2005 PMC : PMC547285
Transposon mutagenesis of a clinical isolate of Morganella morganii, G1492 (tigecycline MIC of 4 microg/ml), yielded two insertion knockout mutants for which tigecycline MICs were 0.03 microg/ml. Transposon insertions mapped to acrA, which is constitutively overexpressed in G1492, suggesting a role of the AcrAB efflux pump in decreased susceptibility to tigecycline in M. morganii.
( 2011 )
Structure of the extended-spectrum �]-lactamase TEM-72 inhibited by citrate.
PMID : 21393831 : DOI : 10.1107/S1744309110054680 PMC : PMC3053151
TEM-72, a class A �]-lactamase identified in isolates of Enterobacteriaceae, is a quadruple mutant of TEM-1 (Q39K, M182T, G238S and E240K) and shows extended-spectrum �]-lactamase (ESBL) properties arising from the G238S and E240K substitutions. Although many structures of TEM variants have been published, they do not include an enzyme with the simultaneous presence of both of the ESBL-conferring G238S and E240K substitutions. Furthermore, the structure shows the presence of a citrate anion bound to the TEM-72 active site, where it interacts with all of the conserved residues of class A �]-lactamases. The present structure supports the use of polycarboxylates as a scaffold for the design of broad-spectrum inhibitors of serine �]-lactamases.
( 2011 )
Phylogenetic analysis of the genera Proteus, Morganella and Providencia by comparison of rpoB gene sequences of type and clinical strains suggests the reclassification of Proteus myxofaciens in a new genus, Cosenzaea gen. nov., as Cosenzaea myxofaciens comb. nov.
PMID : 20709916 : DOI : 10.1099/ijs.0.021964-0
Phylogenetic analysis of partial rpoB gene sequences of type and clinical strains belonging to different 16S rRNA gene-fingerprinting ribogroups within 11 species of enterobacteria of the genera Proteus, Morganella and Providencia was performed and allowed the definition of rpoB clades, supported by high bootstrap values and confirmed by ?2.5 % nucleotide divergence. None of the resulting clades included strains belonging to different species and the majority of the species were confirmed as discrete and homogeneous. However, more than one distinct rpoB clade could be defined among strains belonging to the species Proteus vulgaris (two clades), Providencia alcalifaciens (two clades) and Providencia rettgeri (three clades), suggesting that some strains represent novel species according to the genotypes outlined by rpoB gene sequence analysis. Percentage differences between the rpoB gene sequence of the type strain of Proteus myxofaciens and other members of the same genus (17.3-18.9 %) were similar to those calculated amongst strains of the genus Providencia (16.4-18.7 %), suggesting a genetic distance at the genus-level between Proteus myxofaciens and the rest of the Proteus-Providencia group. Proteus myxofaciens therefore represents a member of a new genus, for which the name Cosenzaea gen. nov., is proposed.
( 2008 )
Tetracycline-resistance genes in gram-negative isolates from estuarine waters.
PMID : 19120920 : DOI : 10.1111/j.1472-765X.2008.02452.x
To investigate the diversity and dissemination of tetracycline resistance genes in isolates from estuarine waters. Forty-two out of 164 multi-resistant isolates previously obtained were resistant or less-susceptible to tetracycline, as evaluated by the disc diffusion method. Minimal inhibitory concentration for resistant bacteria ranged from 16 to 256 mg l(-1). Screening of tet genes by polymerase chain reaction showed that 88% of the isolates carried at least one of the genes tested, namely tet(A) (present in 13 isolates), tet(B) (present in 13 isolates), tet(C) (present in 3 isolates), tet(D) (present in 1 isolate), tet(E) (present in 6 isolates) and tet(M) (present in 1 isolate). One isolate carried tet(A) and tet(M). To our knowledge, this study presents the first description of a tet(D) gene in Morganella morganii. Hybridization revealed that tet genes were plasmid-located in 31% of the isolates. Those isolates were included as donors in conjugation experiments and 38% transferred tetracycline resistance. A considerable diversity of tet genes was detected in the estuary. Frequently, these genes were associated with plasmids and could be transferred to Escherichia coli. The results presented provide further evidence of the role played by estuarine reservoirs in antibiotic resistance maintenance and dissemination.
( 2008 )
Occurrence of mecA in nonstaphylococcal pathogens in surface waters.
PMID : 18845826 : DOI : 10.1128/JCM.01035-08 PMC : PMC2576628
de las Rivas B,
( 2007 )
Gene organization of the ornithine decarboxylase-encoding region in Morganella morganii.
PMID : 17578420 : DOI : 10.1111/j.1365-2672.2006.03188.x
The production of putrescine is a relevant property related to food quality and safety. Morganella morganii is responsible for putrescine production in fresh fish decomposition. The aim of this study was to gain deeper insights into the genetic determinants for putrescine production in M. morganii. The 6972 bp DNA region showed the presence of three complete and two partial open reading frames all transcribed in the same direction. The second and third genes putatively coded for an ornithine decarboxylase (SpeF) and a putrescine-ornithine antiporter (PotE), respectively, and constituted an operon. The speF gene has been expressed in Escherichia coli HT414, an ornithine decarboxylase defective mutant, resulting in ornithine decarboxylase activity. The genetic organization of the SpeF-PotE-encoding region in M. morganii is different to that of E. coli and several Salmonella species. The speF gene cloned from M. morganii encodes a functional ornithine decarboxylase involved in putrescine production. Phylogenetic tree based on 16S rDNA showed that ornithine decarboxylase activity is not related to a specific phylogenetic tree branch in Enterobacteriaceae. The identification of the DNA region involved in putrescine production in M. morganii will allow additional research on their induction and regulation in order to minimize putrescine production in foods.
( 2017 )
Identification of new bacteria harboring qnrS and aac(6')-Ib/cr and mutations possibly involved in fluoroquinolone resistance in raw sewage and activated sludge samples from a full-scale WWTP.
PMID : 27984803 : DOI : 10.1016/j.watres.2016.11.056
Wastewater treatment plants (WWTPs) harbor bacteria and antimicrobial resistance genes, favoring gene exchange events and resistance dissemination. Here, a culture-based and metagenomic survey of qnrA, qnrB, qnrS, and aac(6')-Ib genes from raw sewage (RS) and activated sludge (AS) of a full-scale municipal WWTP was performed. A total of 96 bacterial isolates were recovered from nalidixic acid-enrichment cultures. Bacteria harboring the aac(6')-Ib gene predominated in RS, whereas qnrS-positive isolates were specific to AS. Novel qnrS- and aac(6')-Ib-cr positive species were identified: Morganella morganii, Providencia rettgeri, and Pseudomonas guangdongensis (qnrS), and Alcaligenes faecalis and P. rettgeri (aac(6')-Ib-cr). Analysis of qnrS and aac(6')-Ib sequences from isolates and clone libraries suggested that the diversity of qnrS is wider than that of aac(6')-Ib. A large number of amino acid mutations were observed in the QnrS and AAC(6')-Ib proteins at previously undetected positions, whose structural implications are not clear. An accumulation of mutations at the C72, Q73, L74, A75 and M76 positions of QnrS, and D181 of AAC(6')-Ib might be important for resistance. These findings add significant information on bacteria harboring qnrS and aac(6')-Ib genes, and the presence of novel mutations that may eventually emerge in clinical isolates.
( 1989 )
Isolation and analysis of the C-terminal signal directing export of Escherichia coli hemolysin protein across both bacterial membranes.
PMID : 2656259 : PMC : PMC400846
We have studied the C-terminal signal which directs the complete export of the 1024-amino-acid hemolysin protein (HlyA) of Escherichia coli across both bacterial membranes into the surrounding medium. Isolation and sequencing of homologous hlyA genes from the related bacteria Proteus vulgaris and Morganella morganii revealed high primary sequence divergence in the three HlyA C-termini and highlighted within the extreme terminal 53 amino acids the conservation of three contiguous sequences, a potential 18-amino-acid amphiphilic alpha-helix, a cluster of charged residues, and a weakly hydrophobic terminal sequence rich in hydroxylated residues. Fusion of the C-terminal 53 amino acid sequence to non-exported truncated Hly A directed wild-type export but export was radically reduced following independent disruption or progressive truncation of the three C-terminal features by in-frame deletion and the introduction of translation stop codons within the 3' hlyA sequence. The data indicate that the HlyA C-terminal export signal comprises multiple components and suggest possible analogies with the mitochondrial import signal. Hemolysis assays and immunoblotting confirmed the intracellular accumulation of non-exported HlyA proteins and supported the view that export proceeds without a periplasmic intermediate. Comparison of cytoplasmic and extracellular forms of an independently exported extreme C-terminal 194 residue peptide showed that the signal was not removed during export.
( 2015 )
War wound treatment complications due to transfer of an IncN plasmid harboring bla(OXA-181) from Morganella morganii to CTX-M-27-producing sequence type 131 Escherichia coli.
PMID : 25870058 : DOI : 10.1128/AAC.04442-14 PMC : PMC4432134
A 22-year-old male developed a recurrent sacral abscess associated with embedded shrapnel following a blast injury. Cultures grew extended-spectrum �]-lactamase (ESBL)-producing, carbapenem-susceptible Escherichia coli. Ertapenem was administered, but the infection recurred after each course of antibiotics. Initial surgical interventions were unsuccessful, and subsequent cultures yielded E. coli and Morganella morganii, both nonsusceptible to carbapenems. The isolates were Carba NP test negative, gave ambiguous results with the modified Hodge test, and amplified the bla(OXA48)-like gene by real-time PCR. All E. coli isolates were sequence type 131 (ST131), carried nine resistance genes (including bla(CTX-M-27)) on an IncF plasmid, and were identical by genome sequencing, except for 150 kb of plasmid DNA in carbapenem-nonsusceptible isolates only. Sixty kilobases of this was shared by M. morganii and represented an IncN plasmid harboring bla(OXA-181). In M. morganii, the gene was flanked by IS3000 and ISKpn19, but in all but one of the E. coli isolates containing bla(OXA-181), a second copy of ISKpn19 had inserted adjacent to IS3000. To the best of our knowledge, this is the first report of bla(OXA-181) in the virulent ST131 clonal group and carried by the promiscuous IncN family of plasmids. The tendency of M. morganii to have high MICs of imipenem, a bla(OXA-181) substrate profile that includes penicillins but not extended-spectrum cephalosporins, and weak carbapenemase activity almost resulted in the presence of bla(OXA-181) being overlooked. We highlight the importance of surveillance for carbapenem resistance in all species, even those with intrinsic resistances, and the value of advanced molecular techniques in detecting subtle genetic changes.
( 2015 )
Development of a real-time PCR method coupled with a selective pre-enrichment step for quantification of Morganella morganii and Morganella psychrotolerans in fish products.
PMID : 25791250 : DOI : 10.1016/j.ijfoodmicro.2015.03.005
Histamine fish poisoning is common and due to toxic concentrations of histamine often produced by Gram-negative bacteria in fin-fish products with a high content of the free amino acid histidine. The genus Morganella includes two species previously reported to cause incidents of histamine fish poisoning. Morganella morganii and Morganella psychrotolerans are both strong producer of histamine. However, little is known about the occurrence and critical stages for fish contamination with these bacteria. To elucidate contamination routes of Morganella, specific real-time quantitative PCR (RTi qPCR) methods for quantification of M. morganii and M. psychrotolerans have been developed. Selective primers amplified a 110 bp region of the vasD gene for M. psychrotolerans and a 171 bp region of the galactokinase gene for M. morganii. These primer-sets showed high specificity as demonstrated by using purified DNA from 23 other histamine producing bacteria and 26 isolates with no or limited histamine production. The efficiency of the qPCR reactions on artificially contaminated fish samples were 100.8% and 96.3% respectively. The limit of quantification (LOQ) without enrichment was 4 log CFU/g. A quantitative enrichment step with a selective medium was included and improved the sensitivity of the methods to a LOQ of below 50 CFU/g in seafood. RTi qPCR methods with or without enrichment were evaluated for enumeration of Morganella species in naturally contaminated fresh fish and lightly preserved seafood from Denmark. These new methods will contribute to a better understanding of the occurrence and histamine production by Morganella species in fish products, information that is essential to reduce the unacceptably high frequency of histamine fish poisoning.
( 2014 )
High diversity of beta-lactamases in the General Hospital Vienna verified by whole genome sequencing and statistical analysis.
PMID : 25159028 : DOI : 10.1016/j.meegid.2014.08.014
The detailed analysis of antibiotic resistance mechanisms is essential for understanding the underlying evolutionary processes, the implementation of appropriate intervention strategies and to guarantee efficient treatment options. In the present study, 110 �]-lactam-resistant, clinical isolates of Enterobacteriaceae sampled in 2011 in one of Europe's largest hospitals, the General Hospital Vienna, were screened for the presence of 31 �]-lactamase genes. Twenty of those isolates were selected for whole genome sequencing (WGS). In addition, the number of �]-lactamase genes was estimated using biostatistical models. The carbapenemase genes blaKPC-2, blaKPC-3, and blaVIM-4 were identified in carbapenem-resistant and intermediate susceptible isolates, blaOXA-72 in an extended-spectrum �]-lactamase (ESBL)-positive one. Furthermore, the observed high prevalence of the acquired blaDHA-1 and blaCMY AmpC �]-lactamase genes (70%) in phenotypically AmpC-positive isolates is alarming due to their capability to become carbapenem-resistant upon changes in membrane permeability. The statistical analyses revealed that approximately 55% of all �]-lactamase genes present in the General Hospital Vienna were detected by this study. In summary, this work gives a very detailed picture on the disseminated �]-lactamases and other resistance genes in one of Europe's largest hospitals.
( 2014 )
Structure of the membrane protein MerF, a bacterial mercury transporter, improved by the inclusion of chemical shift anisotropy constraints.
PMID : 25103921 : DOI : 10.1007/s10858-014-9852-0 PMC : PMC4154067
( 2013 )
The structure of the mercury transporter MerF in phospholipid bilayers: a large conformational rearrangement results from N-terminal truncation.
PMID : 23763519 : DOI : 10.1021/ja4042115 PMC : PMC3763827
The three-dimensional structure of the 81-residue mercury transporter MerF determined in liquid crystalline phospholipid bilayers under physiological conditions by Rotationally Aligned (RA) solid-state NMR has two long helices, which extend well beyond the bilayer, with a well-defined interhelical loop. Truncation of the N-terminal 12 residues, which are mobile and unstructured when the protein is solubilized in micelles, results in a large structural rearrangement of the protein in bilayers. In the full-length protein, the N-terminal helix is aligned nearly parallel to the membrane normal and forms an extension of the first transmembrane helix. By contrast, this helix adopts a perpendicular orientation in the truncated protein. The close spatial proximity of the two Cys-containing metal binding sites in the three-dimensional structure of full-length MerF provides insights into possible transport mechanisms. These results demonstrate that major changes in protein structure can result from differences in amino acid sequence (e.g., full-length vs truncated proteins) as well as the use of a non-native membrane mimetic environment (e.g., micelles) vs liquid crystalline phospholipid bilayers. They provide further evidence of the importance of studying unmodified membrane proteins in near-native bilayer environments in order to obtain accurate structures that can be related to their functions.
( 2012 )
Structure determination of a membrane protein in proteoliposomes.
PMID : 22217388 : DOI : 10.1021/ja209464f PMC : PMC3272072
An NMR method for determining the three-dimensional structures of membrane proteins in proteoliposomes is demonstrated by determining the structure of MerFt, the 60-residue helix-loop-helix integral membrane core of the 81-residue mercury transporter MerF. The method merges elements of oriented sample (OS) solid-state NMR and magic angle spinning (MAS) solid-state NMR techniques to measure orientation restraints relative to a single external axis (the bilayer normal) from individual residues in a uniformly (13)C/(15)N labeled protein in unoriented liquid crystalline phospholipid bilayers. The method relies on the fast (>10(5) Hz) rotational diffusion of membrane proteins in bilayers to average the static chemical shift anisotropy and heteronuclear dipole-dipole coupling powder patterns to axially symmetric powder patterns with reduced frequency spans. The frequency associated with the parallel edge of such motionally averaged powder patterns is exactly the same as that measured from the single line resonance in the spectrum of a stationary sample that is macroscopically aligned parallel to the direction of the applied magnetic field. All data are collected on unoriented samples undergoing MAS. Averaging of the homonuclear (13)C/(13)C dipolar couplings, by MAS of the sample, enables the use of uniformly (13)C/(15)N labeled proteins, which provides enhanced sensitivity through direct (13)C detection as well as the use of multidimensional MAS solid-state NMR methods for resolving and assigning resonances. The unique feature of this method is the measurement of orientation restraints that enable the protein structure and orientation to be determined in unoriented proteoliposomes.
( 2012 )
Description and plasmid characterization of qnrD determinants in Proteus mirabilis and Morganella morganii.
PMID : 22192340 : DOI : 10.1111/j.1469-0691.2011.03728.x
We investigated the presence of qnrC and qnrD among 756 non-replicate Enterobacteriaceae isolated in Italy, selected for being non-susceptible to fluoroquinolones and/or resistant to third-generation cephalosporins. Four Proteus mirabilis and one Morganella morganii (0.66% of the total) presented a qnrD gene, located in a 2687-base-pair plasmid that was entirely sequenced. The plasmid is un-typable, and contains no known coding region other than qnrD. That the qnrD gene was found in four unrelated P. mirabilis and in one M. morganii isolate might suggest a frequent association of this gene with the tribe Proteeae.
( 1997 )
Cloning and sequencing of the gene encoding the AmpC beta-lactamase of Morganella morganii.
PMID : 9066104 : DOI : 10.1111/j.1574-6968.1997.tb10260.x
The chromosomal beta-lactamase gene of a clinical isolate of Morganella morganii was cloned in Escherichia coli and sequenced. The beta-lactamase had a pI of 7.4 and conferred a typical AmpC susceptibility pattern. The insert obtained was found to encode a protein of 379 amino acids. Its deduced amino acid sequence revealed it to be a class C beta-lactamase: 39-56% identity with chromosomal AmpC beta-lactamases of Serratia marcescens, Yersinia enterocolitica, Citrobacter freundii, Enterobacter cloacae and Escherichia coli; and 37-56% identity with plasmid-mediated beta-lactamases (MOX-1, CMY-1, FOX-1, ACT-1, LAT-1, BIL-1 and CMY-2). The ampC gene was linked to a gene only part of which (450 bp) was cloned homologous to the regulatory ampR genes of chromosomal class C beta-lactamases.
( 1996 )
A bifunctional urease enhances survival of pathogenic Yersinia enterocolitica and Morganella morganii at low pH.
PMID : 8932305 : DOI : 10.1128/jb.178.22.6487-6495.1996 PMC : PMC178535
To infect a susceptible host, the gastrointestinal pathogen Yersinia enterocolitica must survive passage through the acid environment of the stomach. In this study, we showed that Y. enterocolitica serotype O8 survives buffered acidic conditions as low as pH 1.5 for long periods of time provided urea is available. Acid tolerance required an unusual cytoplasmically located urease that was activated 780-fold by low-pH conditions. Acid tolerance of Helicobacter species has also been attributed to urease activity, but in that case urease was not specifically activated by low-pH conditions. A ure mutant strain of Y. enterocolitica was constructed which was hypersensitive to acidic conditions when urea was available and, unlike the parental strain, was unable to grow when urea was the sole nitrogen source. Examination of other urease-producing gram-negative bacteria indicated that Morganella morganii survives in acidic conditions but Escherichia coli 1021, Klebsiella pneumoniae, Proteus mirabilis, Providencia stuartii, and Pseudomonas aeruginosa do not. Consistent with these results, biochemical evidence demonstrated that Y. enterocolitica and M. morganii ureases were activated in vitro by low pH with an unusually low activity optimum of pH 5.5. In whole cells activation occurred as medium values decreased below pH 3.0 for Y. enterocolitica and pH 5.5 for M. morganii, suggesting that in vivo activation occurs as a result of cytoplasmic acidification. DNA sequence analysis of portions of the M. morganii ure locus showed that the predicted primary structure of the enzyme structural subunits is most similar to those of Y. enterocolitica urease. One region of similarity between these two ureases located near the active site is distinct from most other ureases but is present in the urease of Lactobacillus fermentum. This region of similarity may be responsible for the unique properties of the Y. enterocolitica and M. morganii ureases since the L. fermentum urease also has been shown to have a low pH optimum for activity.
( N/A )
Regulation of the Escherichia coli S10 ribosomal protein operon by heterologous L4 ribosomal proteins.
PMID : 8722027 :
We have cloned the L4 ribosomal protein genes from Morganella morganii and Haemophilus influenza. The sequences of these genes were compared with published sequences for Escherichia coli, Yersinia pseudotuberculosis, and Bacillus stearothermophilus. All five of these L4 genes were expressed in E. coli and shown to function as repressors of both transcription and translation of the E. coli S10 operon. Possible implications for regulation of r-protein synthesis in species other E. coli are discussed.
( 1994 )
Characterization and sequence of PhoC, the principal phosphate-irrepressible acid phosphatase of Morganella morganii.
PMID : 8081499 : DOI : 10.1099/00221287-140-6-1341
Phosphatase activities were investigated in Morganella morganii, which is one of the few enterobacterial species producing high-level phosphate-irrepressible acid phosphatase activity (HPAP phenotype), and the gene encoding the major phosphate-irrepressible acid phosphatase was cloned, sequenced, and its product characterized. Using p-nitrophenyl phosphate as substrate, Morganella produced a major phosphate-irrepressible acid phosphatase (named PhoC) which is associated with the HPAP phenotype, a minor phosphate-irrepressible acid phosphatase, and a phosphate-repressible alkaline phosphatase. The presence of the PhoC activity prevented induction of alkaline phosphatase when a PhoC-hydrolysable organic phosphate ester, such as glycerol 2-phosphate, was the sole phosphate source. PhoC is a secreted nonspecific acid phosphatase apparently composed of four 25 kDa polypeptide subunits. The enzyme is resistant to EDTA, P(i), fluoride and tartrate. The M. morganii PhoC showed 84.6% amino acid sequence identity to the PhoN nonspecific acid phosphatase of Providencia stuartii, 45.3% to the PhoN nonspecific acid phosphatase of Salmonella typhimurium, and 37.8% to the principal acid phosphatase (PhoC) of Zymomonas mobilis. Comparison of sequence data and of regulation of these enzymes suggested a different phylogeny of members of this gene family within the Enterobacteriaceae.
( 1995 )
Purification and properties of crystalline 3-methylaspartase from two facultative anaerobes, Citrobacter sp. strain YG-0504 and Morganella morganii strain YG-0601.
PMID : 7765982 :
3-Methylaspartase (3-methylaspartate ammonia-lyase, EC 22.214.171.124) from two facultative anaerobes from soil, Citrobacter sp. strain YG-0504 and Morganella morganii strain YG-0601, were purified and crystallized from their crude extracts. Both of the Citrobacter and Morganella enzymes appeared to be a dimer of subunits of M(r) 40,000 and 44,000, respectively. The enzymes had similar enzymological properties: optimum pH for the deamination reaction of (2S,3S)-3-methylaspartic acid, substrate specificity, inhibitor, divalent and monovalent cation requirement, and N-terminal amino acid sequence homology. However, some differences were detected in pH and temperature stability, optimum pH for the amination reaction of mesaconic acid, optimum temperature, specific activity, and stability during electrophoresis. Both enzymes had similar enzymological properties to the known 3-methylaspartase from an obligate anaerobic bacterium, Clostridium tetanomorphum H1, except kinetic constants and substrate specificities.
( 1995 )
Cloning and characterization of the NapA acid phosphatase/phosphotransferase of Morganella morganii: identification of a new family of bacterial acid-phosphatase-encoding genes.
PMID : 7894706 : DOI : 10.1099/00221287-141-1-147
The gene encoding a minor phosphate-irrepressible acid phosphatase (named NapA) of Morganella morganii was cloned and sequenced, and its product characterized. NapA is a secreted acid phosphatase composed of four 27 kDa polypeptide subunits. The enzyme is active on several organic phosphate monoesters but not on diesters, and is also endowed with transphosphorylating activity from organic phosphoric acid esters to nucleosides and other compounds with free hydroxyl groups. Its activity is inhibited by EDTA, inorganic phosphate, nucleosides and Ca2+, but not by fluoride or tartrate, and is enhanced by Mg2+, Co2+ and Zn2+. At the sequence level, the NapA enzyme did not show similarities to any other sequenced bacterial phosphatases. However, a search for homologous genes in sequence databases allowed identification of two open reading frames located within sequenced regions of the Escherichia coli and Proteus mirabilis genomes respectively, encoding proteins of unknown function which are highly homologous to the Morganella enzyme. Moreover, the properties of the NapA enzyme are very similar to those reported for the periplasmic nonspecific acid phosphatase II of Salmonella typhimurium (for which no sequence data are available). These data point to the existence of a new family of bacterial acid phosphatases, which we propose designating class B bacterial acid phosphatases.
( 1983 )
Comparison of the lipoprotein gene among the enterobacteriaceae. DNA sequence of Morganella morganii lipoprotein gene and its expression in Escherichia coli.
PMID : 6305973 :
A DNA sequence of 532 base pairs encompassing the entire Morganella morganii lipoprotein gene (lpp) was determined. Sequence comparisons of the M. morganii lpp gene with the lpp genes from Escherichia coli, Serratia marcescens, and Erwinia amylovora reveal that the M. morganii lpp gene is more distantly related to the E. coli lpp gene than any of the other lpp genes examined. Between the E. coli and M. morganii lpp genes, the following homologies were found: 44% in the promoter region (bases, -45 to -1), 88% in the 5'-end untranslated region of the mRNA, 58% in the signal sequence coding region, 75% in the coding region for the first 51 and 43% for the last 7 amino acid residues. Upstream of the promoter region and downstream of the termination codon, there are extensive insertions, deletions, and base substitutions. In spite of the differences in the DNA sequences, the lipoprotein structure was found to be highly conserved except for the carboxyl-terminal sequence of 7 amino residues. The coding region of the M. morganii lpp gene including the signal sequence was inserted into an expression cloning vector so that the production of the M. morganii lipoprotein could be induced in E. coli by a lac inducer, isopropyl-beta-D-thioglactoside. It was found that when induced, the M. morganii prolipoprotein was apparently secreted normally across the E. coli cytoplasmic membrane, modified with glycerol and palmitic acid, processed to the mature lipoprotein, and assembled in the E. coli outer membrane. The bound form covalently linked to the peptidoglycan was also found.
( 1985 )
Inducible xylitol dehydrogenases in enteric bacteria.
PMID : 3886639 : PMC : PMC218933
Morganella morganii ATCC 25829, Providencia stuartii ATCC 25827, Serratia marcescens ATCC 13880, and Erwinia sp. strain 4D2P were found to induce a xylitol dehydrogenase when grown on a xylitol-containing medium. The xylitol dehydrogenases were partially purified from the four strains, and those from M. morganii ATCC 25829, P. stuartii ATCC 25827, and S. marcescens ATCC 13880 were all found to oxidize xylitol to D-xylulose. These three enzymes had KmS for xylitol of 7.1 to 16.4 mM and molecular weights ranging from 130,000 to 155,000. In contrast, the xylitol dehydrogenase from Erwinia sp. strain 4D2P oxidized xylitol at the C-4 position to produce L-xylulose, had a Km for xylitol of 72 mM, and had a molecular weight of 102,000.
( 1986 )
Pyridoxal 5'-phosphate-dependent histidine decarboxylase. Inactivation by alpha-fluoromethylhistidine and comparative sequences at the inhibitor- and coenzyme-binding sites.
PMID : 3733745 :
Pyridoxal phosphate-dependent histidine decarboxylase from Morganella morganii AM-15 was inactivated by (S)-alpha-fluoromethylhistidine by a pseudo first-order reaction, with KI and k inact values of 0.1 mM and 32.2 min-1, respectively, and was most efficient at pH 6.5-7.0. Both L-histidine and the competitive inhibitor, L-histidine methyl ester, protected against inactivation. The apoenzyme was not inactivated. These findings indicate that inhibition is a mechanism-based process. Under optimal conditions a single molecule of alpha-fluoromethylhistidine inactivates one enzyme subunit, indicating that no escaping side reaction occurs during the inactivation process. The bound inactivator is not released by dialysis of the native protein but is released upon denaturation by heat or urea. This released product was not fully characterized, but it contains the tritium of ring-labeled alpha-fluoromethyl-[3H]histidine, exhibits the spectral properties of a 3-hydroxypyridine derivative, and does not yield any amino acids on hydrolysis. The label was much more stable following borohydride reduction of the inactivated protein, and a tryptic peptide containing the modified residue was isolated. Sequencing of this peptide and the corresponding peptide from the native enzyme revealed that the inactivator binds to a serine residue of the holoenzyme. Two P-pyridoxyl peptides from tryptic or CNBr digests of the NaBH4-reduced enzyme were also isolated. Sequence and compositional data obtained with these peptides showed that the serine residue to which the inhibitor binds is not near the lysine residue that binds pyridoxal-P in the primary sequence of the protein, although the two residues must be near one another in the three-dimensional structure to account for these results. A speculative mechanism for inactivation, consistent with the experimental findings, is presented.
( 2019 )
A novel cfr-carrying Tn7 transposon derivative characterized in Morganella morganii of swine origin in China.
PMID : 30508103 : DOI : 10.1093/jac/dky494
To characterize the presence and genetic environment of the multiresistance gene cfr in bacterial isolates from a swine farm. A total of 97 bacterial isolates, recovered from 32 faecal swabs obtained on one farm, were tested for the presence of the cfr gene by PCR. Species identification of the one cfr-positive strain was conducted using the BD PhoenixTM 100 Automated Microbiology System. Susceptibility testing was carried out by broth microdilution. The genetic environment of the cfr gene was analysed by WGS. The Morganella morganii isolate BCMM24 was the only cfr-positive strain. The cfr gene, as well as 15 other resistance genes, is located on a novel 111238 bp transposon derived from Tn7, designated as Tn6451, which comprises various genetic materials including a novel class 1 integron with five gene cassettes. The cfr-containing region consists of a novel genetic structure IS26-cfr-�GTn554 tnpB-�GTn3 family tnpA-IS26, differing from previous reports. Two-step PCR results show that the structure can be looped out and that Tn6451 cannot be excised from the chromosome. To the best of our knowledge, we report the cfr gene in M. morganii for the first time. The cfr gene and 15 other resistance genes are located on a novel Tn7 transposon derivative, suggesting that the Tn7 transposon may act as a reservoir for various antimicrobial resistance genes and more Tn7 derivatives carrying multiple resistance genes are likely to be discovered in Gram-negative bacteria of both animal and human origin.
( 1986 )
Pyridoxal 5'-phosphate-dependent histidine decarboxylase. Nucleotide sequence of the hdc gene and the corresponding amino acid sequence.
PMID : 3015950 :
The nucleotide sequence of a 1.3-kilobase NaeI fragment from Morganella morganii AM-15 that contains the gene for histidine decarboxylase has been determined. The gene was initially identified among total chromosomal digests using a mixed sequence oligonucleotide probe corresponding to amino acids 11-16 of histidine decarboxylase and then cloned on a 5.5-kilobase PstI fragment. The structural gene contains 1131 nucleotides and encodes 377 amino acids with the sequence: (sequence: in text). The independently determined NH2-terminal sequence of this enzyme (Tanase, S., Guirard, B. M., and Snell, E. E. (1985) J. Biol. Chem. 260, 6738-6746) and the amino acid sequences of two tryptic peptides reported in the accompanying paper (Hayashi, H., Tanase, S., and Snell, E. E. (1986) J. Biol. Chem. 261, 11003-11009) are localized in the sequence presented here; the lysine that binds pyridoxal phosphate is situated at residue 232, whereas the serine that binds the adduct formed between pyridoxal phosphate and the inhibitor alpha-fluoromethylhistidine is positioned at residue 322.
( 2013 )
Prevalence and plasmid characterization of the qnrD determinant in Enterobacteriaceae isolated from animals, retail meat products, and humans.
PMID : 23557071 : DOI : 10.1089/mdr.2012.0146
qnrD, unlike other qnr genes, is mainly located on small nonconjugative plasmids. We investigated the presence of qnrD among 1,373 Enterobacteriaceae isolates in China. Twelve qnrD-positive strains were detected, and all were nonsusceptible to fluoroquinolones. The complete sequence of plasmids showed that the qnrD determinants were located on two plasmids with a respective size of ~4.2 and 2.7 k-bp. Interestingly, the identification of qnrD in this study revealed the highest prevalence of Proteeae among Enterobacteriaceae identified.
( 2012 )
Emergence of New Delhi metallo-�]-lactamase in Jerusalem, Israel.
PMID : 22951226 : DOI : 10.1016/j.ijantimicag.2012.07.011
( 1990 )
Morganella morganii urease: purification, characterization, and isolation of gene sequences.
PMID : 2345135 : DOI : 10.1128/jb.172.6.3073-3080.1990 PMC : PMC209110
Morganella morganii, a very common cause of catheter-associated bacteriuria, was previously classified with the genus Proteus on the basis of urease production. M. morganii constitutively synthesizes a urease distinct from that of other uropathogens. The enzyme, purified 175-fold by passage through DEAE-Sepharose, phenyl-Sepharose, Mono-Q, and Superose 6 chromatography resins, was found to have a native molecular size of 590 kilodaltons and was composed of three distinct subunits with apparent molecular sizes of 63, 15, and 6 kilodaltons, respectively. Amino-terminal analysis of the subunit polypeptides revealed a high degree of conservation of amino acid sequence between jack bean and Proteus mirabilis ureases. Km for urea equalled 0.8 mM. Antiserum prepared against purified enzyme inhibited activity by 43% at a 1:2 dilution after 1 h of incubation. All urease activity was immunoprecipitated from cytosol by a 1:16 dilution. Antiserum did not precipitate ureases of other species except for one Providencia rettgeri strain but did recognize the large subunits of ureases of Providencia and Proteus species on Western blots (immunoblots). Thirteen urease-positive cosmid clones of Morganella chromosomal DNA shared a 3.5-kilobase (kb) BamHI fragment. Urease gene sequences were localized to a 7.1-kb EcoRI-SalI fragment. Tn5 mutagenesis revealed that between 3.3 and 6.6 kb of DNA were necessary for enzyme activity. A Morganella urease DNA probe did not hybridize with gene sequences of other species tested. Morganella urease antiserum recognized identical subunit polypeptides on Western blots of cytosol from the wild-type strain and Escherichia coli bearing the recombinant clone which corresponded to those seen in denatured urease. Although the wild-type strain and recombinant clone produced equal amounts of urease protein, the clone produced less than 1% of the enzyme activity of the wild-type strain.
( 2013 )
New integron gene arrays from multiresistant clinical isolates of members of the Enterobacteriaceae and Pseudomonas aeruginosa from hospitals in Malaysia.
PMID : 23180481 : DOI : 10.1099/jmm.0.053645-0
This study investigated 147 multidrug-resistant Enterobacteriaceae and Pseudomonas aeruginosa isolates from hospitalized patients in Malaysia. Class 1 integrons were the most dominant class identified (45.6%). Three isolates were shown to contain class 2 integrons (2.0%), whilst one isolate harboured both class 1 and 2 integrons. No class 3 integrons were detected in this study. In addition, the sul1 gene was amplified in 35% of isolates and was significantly associated with the presence of integrase genes in an integron structure. RFLP and DNA sequencing analyses revealed the presence of 19 different cassette arrays among the detected integrons. The most common gene cassettes were those encoding resistance towards aminoglycosides (aad) and trimethoprim (dfr). As far as is known, this study is the first to identify integron-carrying cassette arrays such as aadA2-linF, aacC3-cmlA5 and aacA4-catB8-aadA1 in the Malaysian population. Patients' age was demonstrated as a significant risk factor for the acquisition of integrons (P=0.028). Epidemiological typing using PFGE also demonstrated a clonal relationship among isolates carrying identical gene cassettes in Klebsiella pneumoniae and P. aeruginosa but not in Escherichia coli isolates.
( 2013 )
Differential distribution of plasmid-mediated quinolone resistance genes in clinical enterobacteria with unusual phenotypes of quinolone susceptibility from Argentina.
PMID : 23478955 : DOI : 10.1128/AAC.01615-12 PMC : PMC3716127
We studied a collection of 105 clinical enterobacteria with unusual phenotypes of quinolone susceptibility to analyze the occurrence of plasmid-mediated quinolone resistance (PMQR) and oqx genes and their implications for quinolone susceptibility. The oqxA and oqxB genes were found in 31/34 (91%) Klebsiella pneumoniae and 1/3 Klebsiella oxytoca isolates. However, the oqxA- and oqxB-harboring isolates lacking other known quinolone resistance determinants showed wide ranges of susceptibility to nalidixic acid and ciprofloxacin. Sixty of the 105 isolates (57%) harbored at least one PMQR gene [qnrB19, qnrB10, qnrB2, qnrB1, qnrS1, or aac(6')-Ib-cr)], belong to 8 enterobacterial species, and were disseminated throughout the country, and most of them were categorized as susceptible by the current clinical quinolone susceptibility breakpoints. We developed a disk diffusion-based method to improve the phenotypic detection of aac(6')-Ib-cr. The most common PMQR genes in our collection [qnrB19, qnrB10, and aac(6')-Ib-cr] were differentially distributed among enterobacterial species, and two different epidemiological settings were evident. First, the species associated with community-acquired infections (Salmonella spp. and Escherichia coli) mainly harbored qnrB19 (a unique PMQR gene) located in small ColE1-type plasmids that might constitute its natural reservoirs. qnrB19 was not associated with an extended-spectrum �]-lactamase phenotype. Second, the species associated with hospital-acquired infections (Enterobacter spp., Klebsiella spp., and Serratia marcescens) mainly harbored qnrB10 in ISCR1-containing class 1 integrons that may also have aac(6')-Ib-cr as a cassette within the variable region. These two PMQR genes were strongly associated with an extended-spectrum �]-lactamase phenotype. Therefore, this differential distribution of PMQR genes is strongly influenced by their linkage or lack of linkage to integrons.
( 1999 )
Molecular characterization of a ceftazidime-resistant Morganella morganii isolate producing a TEM-10 beta-lactamase.
PMID : 9989337 : PMC : PMC89100
( 1998 )
Molecular characterization of a TEM-21 beta-lactamase in a clinical isolate of Morganella morganii.
PMID : 9687421 : PMC : PMC105882
A clinical isolate of Morganella morganii, with reduced susceptibility to expanded-spectrum cephalosporins and aztreonam, was found to produce an extended-spectrum beta-lactamase with a pI of 6.4. The nucleotide sequence of the encoding gene was that of the gene encoding TEM-21. This is the first molecular characterization of an extended-spectrum beta-lactamase in M. morganii.
( 1998 )
A family of stability determinants in pathogenic bacteria.
PMID : 9829958 : PMC : PMC107735
A novel segregational stability system was identified on plasmid R485, which originates from Morganella morganii. The system is composed of two overlapping genes, stbD and stbE, which potentially encode proteins of 83 and 93 amino acids, respectively. Homologs of the stbDE genes were identified on the enterotoxigenic plasmid P307 from Escherichia coli and on the chromosomes of Vibrio cholerae and Haemophilus influenzae biogroup aegyptius. The former two homologs also promote plasmid stability in E. coli. Furthermore, the stbDE genes share homology with components of the relBEF operon and with the dnaT gene of E. coli. The organization of the stbDE cassette is reminiscent of toxin-antitoxin stability cassettes.
( 1997 )
E. coli translation initiation factor IF2--an extremely conserved protein. Comparative sequence analysis of the infB gene in clinical isolates of E. coli.
PMID : 9428651 : DOI : 10.1016/s0014-5793(97)01472-5
The functionally uncharacterised N-terminal of translation initiation factor IF2 has been found to be extremely variable when comparing different bacterial species. In order to study the intraspecies variability of IF2 the 2670 basepairs nucleotide sequence of the infB gene (encoding IF2) was determined in 10 clinical isolates of E. coli. The N-terminal domains (I, II and III) were completely conserved indicating a specific function of this region of IF2. Only one polymorphic position was found in the deduced 890 amino acid sequence. This Gln/Gly490 is located within the central GTP/GDP-binding domain IV of IF2. The results are further evidence that IF2 from E. coli has reached a highly defined level of structural and functional development.