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Michael R Mulvey

Publications and source records attributed to Michael R Mulvey.

At least 19 recordsLinked to original sources
PubMed2006

The effect of the Salmonella genomic island 1 on in vitro global gene expression in Salmonella enterica serovar Typhimurium LT2.

A Salmonella genomic island 1 (SGI1) isogenic strain pair was constructed using Salmonella enterica serovar Typhimurium LT2 (ST LT2). Real-time quantitative reverse transcriptase PCR revealed detectable mRNA transcripts for all 44 putative ORFs encoded within the SGI1. The highest levels of transcripts observed in SGI1 encoded ORFs were found in genes conferring antibiotic resistance to ampicillin, streptomycin/spectinomycin, and sulphonamides. Abundant mRNA transcripts, relative to gapA, were also noted for one putative regulatory ORF and seven ORFs of unknown function encoded within SGI1, whose products could represent factors contributing to increases in virulence and/or fitness of the organism. DNA microarray analysis revealed the differential expression of known factors that contribute to virulence in many pathogens. Twenty-two chromosomal genes were significantly upregulated in ST LT2 harboring SGI1, which included increased expression of iron and sialic acid utilization genes. Decreased expression was noted for 15 genes in ST LT2 harboring SGI1, including genes involved in chemotaxis and motility. This is the first report examining gene expression within the SGI1, as well as its potential effect on global gene expression, and sets the foundation for future studies involving the effect of SGI1 in other Salmonella spp.

Conjugation, Genetic↗
PubMed2006

Rapid identification of coagulase-negative staphylococci by Fourier transform infrared spectroscopy.

Coagulase-negative staphylococci (CNS), frequently associated with both community-acquired and nosocomial bloodstream infections, must be distinguished from Staphylococcus aureus for clinical purposes. Conventional methods are too laborious and time-consuming and often lack sensitivity to CNS. Fourier transform infrared (FTIR) spectroscopy combined with the use of a universal growth medium (Que-Bact Universal Medium No. 2) and chemometrics was evaluated for its potential as a rapid and simple clinical tool for making this distinction. FTIR spectra of 11 methicillin-sensitive and 11 methicillin-resistant CNS isolates as well as 25 methicillin-sensitive, 47 methicillin-resistant, 34 borderline oxacillin-resistant and 35 glycopeptide intermediate S. aureus isolates were obtained from dried films of stationary-phase cells grown on the universal medium. Principal component analysis (PCA), self-organizing maps, and the K-nearest neighbor algorithm were employed to cluster the different phenotypes based on similarity of their FTIR spectra. PCA of the first-derivative normalized spectral data from a single narrow region (2888-2868 cm(-1)) yielded complete differentiation of CNS from both methicillin-sensitive and methicillin-resistant S. aureus. The rate of correct classification was somewhat reduced, from 100% to 90%, after inclusion of borderline oxacillin-resistant and glycopeptide intermediate S. aureus strains in the data set. Differentiation based on the data in broader spectral regions was much less reliable. The results of this study indicate that with proper spectral region selection, FTIR spectroscopy and cluster analysis may provide a simple and accurate means of CNS species identification.

Algorithms↗
PubMed2006

Salmonella resistant to extended-spectrum cephalosporins: prevalence and epidemiology.

Salmonella resistant to extended-spectrum cephalosporins (ESCs) have emerged worldwide since 1988. By 2004, 43 countries had reported this public health problem. Resistance was mediated by classical extended-spectrum beta-lactamases, plasmid-mediated cephalosporinases, and recently a class A carbapenemase. Of these, CMY-2 is the most widely disseminated enzyme. Salmonella enterica serotype Typhimurium and S. enterica serotype Enteritidis are the most common serovars associated with ESC resistance in human infections. Many outbreaks in humans have been reported, most often among children and neonates. ESC-resistant Salmonella is frequently recovered from animals and food, with poultry as primary food source, suggesting that humans are often infected by these routes.

Animals↗
PubMed2006

The genetics of Salmonella genomic island 1.

Multidrug-resistant Salmonella enterica serovar Typhimurium phage type DT104, resistant to ampicillin, chloramphenicol/florfenicol, streptomycin, sulfonamides, and tetracycline, has disseminated worldwide. The resistance genes reside on the 43-kb Salmonella genomic island 1 (SGI1), which is transferable. Drug-resistant variants of SGI1 have been identified in numerous serotypes. Strains harboring SGI1 may be more virulent and have a tendency to rapidly disseminate.

Animals↗
PubMed2006

VanG-type vancomycin-resistant Enterococcus faecalis strains isolated in Canada.

Enterococcus faecalis G1-0247 (vancomycin MIC, 16 microg/ml) was found to harbor a vanG operon 99% identical to the vanG operon in E. faecalis BM4518. E. faecalis N03-0233 (vancomycin MIC, 16 microg/ml) was found to harbor a novel vanG operon, vanG2, on an element in a different chromosomal location than the vanG-harboring elements in G1-0247 and BM4518.

Anti-Bacterial Agents↗
PubMed2006

Enterococcus gallinarum N04-0414 harbors a VanD-type vancomycin resistance operon and does not contain a D-alanine:D-alanine 2 (ddl2) gene.

Enterococcus gallinarum N04-0414 (MIC for vancomycin, 256 microg/ml) harbored a vanD-type vancomycin resistance operon as well as the intrinsic vanC1 operon. The D-Ala:D-Ala ligase 2 gene (ddl2) was not present in the strain, though it is found downstream of the vanS gene from the vanC operon in E. gallinarum ATCC 49573 and 19 other E. gallinarum strains tested.

Alanine↗
PubMed2006

Single-nucleotide repeat analysis for subtyping Bacillus anthracis isolates.

Single-nucleotide repeats (SNRs) are variable-number tandem repeats that display very high mutation rates. In an outbreak situation, the use of a marker system that exploits regions with very high mutation rates, such as SNRs, allows the differentiation of isolates with extremely low levels of genetic diversity. This report describes the identification and analysis of SNR loci of Bacillus anthracis. SNR loci were selected in silico, and the loci with the highest diversity were used to design and test locus-specific primers against a number of B. anthracis strains with the same multilocus variable-number tandem repeat analysis (MLVA) genotype. SNR markers that allowed strains with the same MLVA genotype to be differentiated from each other were identified. The resulting SNR marker system can be used as a molecular epidemiological tool in a natural outbreak or bioterrorism event, offering the best chance of distinguishing very closely related isolates.

Animals↗
PubMed2005

Increase in ampC promoter strength due to mutations and deletion of the attenuator in a clinical isolate of cefoxitin-resistant Escherichia coli as determined by RT-PCR.

OBJECTIVES: To characterize the mechanism of cefoxitin resistance in clinical isolate Escherichia coli N99-0001. METHODS: Plasmid analysis, PCR for beta-lactamases, and sequencing of the ampC genes was carried out. An RT-PCR method was developed to determine relative ampC expression. RESULTS: Analysis of the ampC promoter region of E. coli N99-0001 revealed a T-->A mutation at -32, a C-->A mutation at -11, an insertion of a T between -20 and -21, and a 28 bp deletion including the entire attenuator. RT-PCR showed that ampC was expressed 140-fold higher in E. coli N99-0001 than in E. coli ATCC 25922. CONCLUSIONS: Cefoxitin resistance in E. coli N99-0001 was due to overexpression of ampC caused by an increase in promoter strength.

Amino Acid Sequence↗
PubMed2005

The Salmonella genomic island 1 is an integrative mobilizable element.

Salmonella genomic island 1 (SGI1) is a genomic island containing an antibiotic resistance gene cluster identified in several Salmonella enterica serovars. The SGI1 antibiotic resistance gene cluster, which is a complex class 1 integron, confers the common multidrug resistance phenotype of epidemic S. enterica Typhimurium DT104. The SGI1 occurrence in S. enterica serovars Typhimurium, Agona, Paratyphi B, Albany, Meleagridis and Newport indicates the horizontal transfer potential of SGI1. Here, we report that SGI1 could be conjugally transferred from S. enterica donor strains to non-SGI1 S. enterica and Escherichia coli recipient strains where it integrated into the recipient chromosome in a site-specific manner. First, an extrachromosomal circular form of SGI1 was identified by PCR which forms through a specific recombination of the left and right ends of the integrated SGI1. Chromosomal excision of SGI1 was found to require SGI1-encoded integrase which presents similarities to the lambdoid integrase family. Second, the conjugal transfer of SGI1 required the presence of a helper plasmid. The conjugative IncC plasmid R55 could thus mobilize in trans SGI1 which was transferred from the donor to the recipient strains. By this way, the conjugal transfer of SGI1 occurred at a frequency of 10(-5)-10(-6) transconjugants per donor. No transconjugants could be obtained for the SGI1 donor lacking the int integrase gene. Third, chromosomal integration of SGI1 occurred via a site-specific recombination between a 18 bp sequence found in the circular form of SGI1 and a similar 18 bp sequence at the 3' end of thdF gene in the S. enterica and E. coli chromosome. SGI1 appeared to be transmissible only in the presence of additional conjugative functions provided in trans. SGI1 can thus be classified within the group of integrative mobilizable elements (IMEs).

Amino Acid Sequence↗
PubMed2005

Molecular characterization of cefoxitin-resistant Escherichia coli from Canadian hospitals.

A study designed to gain baseline information on strains of Escherichia coli displaying resistance to cefoxitin in Canada is described. A total of 29,323 E. coli isolates were screened at 12 participating hospital sites as part of an extended-spectrum beta-lactamase surveillance initiative. A total of 411 clinically significant, nonrepeat isolates displaying reduced susceptibilities to the NCCLS-recommended beta-lactams were submitted to a central laboratory over a 1-year period ending on 30 September 2000. Two hundred thirty-two isolates were identified as resistant to cefoxitin. All cefoxitin-resistant strains were subtyped by pulsed-field gel electrophoresis, and of these, 182 strains revealed a unique fingerprint and 1 strain was untypeable. PCR and sequence analysis of the ampC promoter region revealed 51 different promoter or attenuator variants and 14 wild-type promoters. Three promoter regions were interrupted by insertion elements, two contained IS10 elements, and one contained an IS911 variant. PCR and sequence analysis for the detection of acquired AmpC resistance (by the acquisition of ACT-1/MIR-1, CMY-2, or FOX) revealed that 25 strains contained CMY-2, including 7 of the strains found to have wild-type promoters. The considerable genetic variability in both the strain fingerprint and the promoter region suggests that AmpC-type resistance may emerge spontaneously by mutation of sensitive strains rather than by the spread of strains or plasmids in the hospital setting.

Anti-Bacterial Agents↗
PubMed2005

Identification of a progenitor of the CTX-M-9 group of extended-spectrum beta-lactamases from Kluyvera georgiana isolated in Guyana.

Chromosomal beta-lactamase genes (bla(KLUY)) from six Kluyvera georgiana strains isolated in Guyana were cloned and expressed in Escherichia coli. KLUY-1 exhibited 100% amino acid identity with the extended-spectrum beta-lactamase CTX-M-14. We also show that a 2.7-kb Kluyvera chromosomal region exhibits 99% nucleotide identity to a portion of In60 that includes bla(CTX-M-9).

Amino Acid Sequence↗
PubMed2005

Development of a triplex real-time PCR assay for detection of Panton-Valentine leukocidin toxin genes in clinical isolates of methicillin-resistant Staphylococcus aureus.

Community-associated methicillin-resistant Staphylococcus aureus harboring Panton-Valentine leukocidin (PVL) genes is an emerging pathogen. A novel real-time PCR assay for identification of MRSA isolates containing PVL was developed. The PVL assay was used in a triplex format allowing simultaneous amplification of mecA, nuc, and PVL genes in 614 clinical isolates. This assay facilitates the rapid identification of PVL-positive isolates of MRSA.

Bacterial Toxins↗
PubMed2005

Community-associated methicillin-resistant Staphylococcus aureus, Canada.

A total of 184 methicillin-resistant Staphylococcus aureus (MRSA) strains were collected from patients who sought treatment primarily for skin and soft tissue infections from January 1, 1999, to March 31, 2002, in east-central Saskatchewan, Canada. Molecular subtyping analysis using pulsed-field gel electrophoresis showed 2 major clusters. Cluster A (n = 55) was composed of a multidrug-resistant MRSA strain associated with a long-term care facility and was similar to the previously reported nosocomial Canadian epidemic strain labeled CMRSA-2. Cluster B (n = 125) was associated with cases identified at community health centers and was indistinguishable from a community-associated (CA)-MRSA strain identified previously in the United States (USA400). Cluster B remained susceptible to a number of classes of antimicrobial agents and harbored the lukF-PV and lukS-PV toxin genes. Over 50% of both clonal groups displayed high-level resistance to mupirocin. This is the first report of the USA400 strain harboring the lukF-PV and lukS-PV toxin genes in Canada.

Adolescent↗
PubMed2004

Enterococcus faecium N03-0072 carries a new VanD-type vancomycin resistance determinant: characterization of the VanD5 operon.

OBJECTIVES: To genotypically characterize the vancomycin resistance mechanism of Enterococcus faecium N03-0072, which was negative by PCR for the currently known van genotypes. METHODS: PCR was used to amplify the entire vancomycin resistance operon and the complete nucleotide sequence was determined by dideoxy cycle sequencing. RESULTS: Analysis revealed a VanD-type operon with 94% nucleotide identity to the VanD4 operon and 90% nucleotide identity to the VanD1/D3 operons. A set of universal primers was designed in order to identify all current vanD variants by PCR. CONCLUSIONS: E. faecium N03-0072 carries a new VanD-type operon, designated VanD5.

Bacterial Proteins↗
PubMed2004

Characterization of a Salmonella enterica serovar Agona strain harbouring a class 1 integron containing novel OXA-type beta-lactamase (blaOXA-53) and 6'-N-aminoglycoside acetyltransferase genes [aac(6')-I30].

OBJECTIVE: To characterize by molecular methods a multidrug-resistant Salmonella enterica serovar Agona (S. enterica Agona) isolated from a hospitalized patient in Rio de Janeiro, Brazil. METHODS: The S. enterica Agona strain was screened by PCR and DNA sequencing for TEM, SHV and CTX-M-type beta-lactamase genes, tet(A), (B), (C) and (D) tetracycline resistance genes, chloramphenicol resistance genes and class 1 integrons. Plasmid characterization was carried out by PCR and Southern hybridization analysis. PCR and PFGE were used to characterize nine other S. enterica Agona strains collected from hospitals in Rio de Janeiro. RESULTS: The study strain was found to harbour a 105 kb plasmid, which contained catA1, bla(TEM-1), a class 1 integron with two novel genes labelled bla(OXA-53) and aac(6')-I30, respectively, and an additional unidentified aminoglycoside resistance gene. A second 53 kb plasmid from the same strain contained tet(D) and bla(SHV-5). OXA-53 was shown to provide reduced susceptibility to ceftazidime, and its activity was inhibited in the presence of clavulanic acid. PFGE analysis of the nine other S. enterica Agona strains revealed two clusters of related strains (78% similarity), and PCR analysis showed that all strains contained the novel integron. CONCLUSION: An S. enterica Agona strain was found to harbour three plasmid-encoded beta-lactamases, one (OXA-53) on a novel class 1 integron that also contains a new aminoglycoside resistance gene, aac(6')-I30. The multidrug resistance plasmids appear to have disseminated to other city hospitals via other S. enterica Agona strains.

Acetyltransferases↗