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PubMed2002

Experimental application of the median-effect principle for in vitro quantification of the combined inhibitory activities of clavulanic acid and imipenem against IRT-4 beta-lactamase.

Inhibitor-resistant TEM beta-lactamases (IRT) have been identified in Enterobacteriaceae which, however, remained susceptible to cephalosporins. We evaluated the combined inhibitory activity of clavulanic acid and imipenem at ratios of 1:1 and 1:3 against IRT-4, using the median effect principle of Chou and Talalay. The combination of the two drugs, which produced a nearly additive effect, meant their concentrations could be lowered 1.3-4.9-fold, while maintaining a 50% inhibitory effect against the IRT-4 in comparison with each drug alone. From a therapeutic point of view, such a combination is not efficient but this method of Chou and Talalay, used for the first time to assay combined inhibitory activity of beta-lactamase inhibitors, could be used to evaluate new molecules and/or strategies to inactivate beta-lactamase.

Anti-Bacterial Agents↗
PubMed1994

Characterization and amino acid sequence of IRT-4, a novel TEM-type enzyme with a decreased susceptibility to beta-lactamase inhibitors.

The clinical isolate Escherichia coli PEY was highly resistant to amoxycillin, ticarcillin and piperacillin associated to beta-lactamase inhibitors such as clavulanic acid, sulbactam, tazobactam and brobactam but susceptible to cephalosporins, aztreonam and imipenem. The susceptibility to mecillinam indicated that this phenotype was not related to hyperproduction of the TEM-1 beta-lactamase. E. coli PEY produced a new plasmid-mediated inhibitor-resistant beta-lactamase of pI 5.2, which was named IRT-4. The determination of the amino acid sequence (Swiss-Prot accession number, P00810) of the purified protein indicated that IRT-4 differed from TEM-1 by two substitutions: Leu for Met-69 (ABL numbering) and Asp for Asn-276. A Met-69-Leu variant of TEM-1, obtained by site-directed mutagenesis, has been described as resistant to clavulanate. The Asp for Asn-276 substitution has not been reported previously. The side chains of Asp-276 and Arg-244 were expected to interact. Determinations of 50% inhibitory concentrations of beta-lactamase inhibitors and substrate profile of IRT-4 suggested that such an ionic bond was implicated in the alteration of the mechanistic process of TEM-1 beta-lactamase.

Amino Acid Sequence↗
PubMed2001

Catalytic and structural properties of IRT-21 beta-lactamase (TEM-77) from a co-amoxiclav-resistant Proteus mirabilis isolate.

Proteus mirabilis strain MAG1, a clinical isolate that is resistant to broad-spectrum penicillins and co-amoxiclav, produces inhibitor-resistant TEM (IRT)-21, a novel mutant of TEM beta-lactamase. This enzyme has a pI of 5.2 and is derived from the bla(TEM-1a) gene ancestor. It contains two major amino acid substitutions specific for co-amoxiclav resistance (Leu-69 for Met and Ser-244 for Arg) that have never been found together previously. The dramatic loss of sensitivity to clavulanic acid, the enhancement of K(m) for all beta-lactams and markedly for ticarcillin, and the decrease in the catalytic efficiency makes IRT-21 comparable to the other IRTs with substitutions at position 244 or double substitutions.

Amoxicillin-Potassium Clavulanate Combination↗
PubMed1995

First characterization of inhibitor-resistant TEM (IRT) beta-lactamases in Klebsiella pneumoniae strains.

Two clinical strains of Klebsiella pneumoniae, TP 01 and TP 02, presented resistance to amoxicillin-clavulanate and were fully susceptible to cephalothin. These strains produced two beta-lactamases, SHV-1 and a TEM enzyme with a pI of 5.2. The previously described changes Arg-244-->Cys and Arg-244-->Ser in IRT-1 and IRT-2 (A. Belaaouaj, C. Lapoumeroulie, M. M. Caniça, G. Vedel, P. Nevot, R. Krishnamoorthy, and G. Paul, FEMS Microbiol. Lett. 120:75-80, 1994) were found in TEM enzymes from the TP 01 and TP 02 strains, respectively. This is the first report of inhibitor-resistant TEM (IRT) in species other than Escherichia coli from the family Enterobacteriaceae.

Amoxicillin↗
PubMed1999

Using IRT variable maps to enrich understanding of rehabilitation data.

One of the benefits of item response theory (IRT) applications in scale development is the greater transparency of resulting scores. This feature allows translation of a total score on a particular scale into a profile of probable item responses with relative ease, for example by using the variable map that often is part of IRT analysis output. Although there have been a few examples in the literature using variable maps to enrich clinical interpretation of individual functional assessment scores, this feature of IRT output has received very limited application in rehabilitation research. The present paper illustrates the application of variable maps to support more in-depth interpretation of functional assessment scores in research and clinical contexts. Two examples are presented from an outcome prediction study conducted during the standardization of a new functional assessment for elementary school students with disabilities, the School Function Assessment. Two different applications are described: creating a dichotomous outcome variable using scores from a continuous scale, and interpreting the meaning of a classification cut-off score identified through Classification and Regression Tree (CART) analysis.

Activities of Daily Living↗
PubMed2004

Use of an IRT-based latent variable model to link different forms of the CES-D from the Health and Retirement Study.

BACKGROUND: The goal of this study was to calibrate depressive symptoms collected using different versions of the Centers for Epidemiologic Studies - Depression (CES-D) instrument in different waves of the Health and Retirement Study (HRS). METHOD: The HRS is a prospective and nationally representative cohort study. This analysis included a sample of HRS participants, adults aged 23-85 years in 1992 who had complete data on depressive symptoms at initial 2- and 4-year follow-up interviews (N= 5,734). Depressive symptoms were assessed with the CES-D. Symptom coverage and response categories varied across study wave. The first wave (1992) used a four-category response, whereas subsequent waves (1994 and 1996) used a two-category response. A structural equations model (SEM) based in Item Response Theory (IRT) was used to calibrate symptoms and generate linked depressive symptom burden scores. RESULTS: Linked depressive symptom burden scores, derived from calibrated symptoms, were distributed similarly across HRS wave. CONCLUSION: Our results demonstrate the applicability of an IRT-based SEM to address a common challenge in prospective studies: changes in the content and context of symptom assessment. Future investigations may make use of our linked syndrome scores to further explore other aspects of depression from a longitudinal perspective.

Adult↗
PubMed2006

Newborn screening for CF in a regional paediatric centre: the psychosocial effects of false-positive IRT results on parents.

Neonatal screening for cystic fibrosis (CF) has been established in Leeds since 1975. The current method is measuring IRT and genotyping. Newborn screening for CF results in a small but significant number of false positives. This study explored the psychosocial reactions to such results in a group of parents (N=21) using semi-structured interviews. Responses were analysed using descriptive statistics and well-validated content analysis. Mothers described a range of emotions during the screening process including anxiety, distress and upset. Waiting for the repeat IRT test results was identified as the most emotionally difficult stage. Discussion focuses on good practice and implications for CF services.

Adult↗
PubMed2000

Using IRT to separate measurement bias from true group differences on homogeneous and heterogeneous scales: an illustration with the MMPI.

The authors present a didactic illustration of how item response theory (IRT) can be used to separate measurement bias from true group differences on homogeneous and heterogeneous scales. Several bias detection methods are illustrated with 12 unidimensional Minnesota Multiphasic Personality Inventory (MMPI) factor scales (Waller, 1999) and the 13 multidimensional MMPI validity and clinical scales. The article begins with a brief review of MMPI bias research and nontechnical reviews of the 2-parameter logistic model (2-PLM) and several IRT-based methods for bias detection. A goal of this article is to demonstrate that homogeneous and heterogeneous scales that are composed of biased items do not necessarily yield biased test scores. To that end, the authors perform differential item- and test-functioning analyses on the MMPI factor, validity, and clinical scales using data from 511 Blacks and 1,277 Whites from the California Youth Authority.

Adolescent↗
PubMed2000

Molecular epidemiology of an outbreak due to IRT-2 beta-lactamase-producing strains of Klebsiella pneumoniae in a geriatric department.

In February 1998, 195 patients in the geriatric department of a French hospital were screened for the presence of co-amoxiclav-resistant Klebsiella pneumoniae. Eleven co-amoxiclav-resistant isolates obtained all produced an identical IRT-2 beta-lactamase. These K. pneumoniae isolates were clonally related and harboured a c. 55 kb non-conjugative plasmid encoding a non-class-1 integron-located blaIRT-2 gene. This study underlines that geriatric departments may be a reservoir for antibiotic-resistant strains and that IRT beta-lactamase-producing strains may be nosocomial pathogens.

Amoxicillin-Potassium Clavulanate Combination↗
PubMed1997

Properties of IRT-14 (TEM-45), a newly characterized mutant of TEM-type beta-lactamases.

IRT-14 (TEM-45) is a new mutant TEM-type beta-lactamase that was isolated from clinical Escherichia coli P37 and that confers resistance to broad-spectrum penicillins with reduced sensitivity to beta-lactamase inhibitors. The MICs of amoxicillin alone and of amoxicillin combined with 2 micrograms of clavulanic acid or 2 micrograms of tazobactam per ml were 4,096, 2,048, and 1,024 micrograms/ml, respectively. The strain was susceptible to cephalosporins, aztreonam, moxalactam, and imipenem. The enzyme was purified to homogeneity, and values of the kinetic parameters Kcat, Km, and Kcat/Km were determined for different substrates. This enzyme, with a pI of 5.2, was found to have reduced affinity for broad-spectrum penicillins and cephalosporins. The values of 50% inhibitory concentrations of clavulanic acid, sulbactam, tazobactam, and brobactam are correlated with the higher KmS for substrates. The resistance of E. coli P37 to mechanism-based inactivators results from a higher level of production of the TEM-derived enzyme due to the G-to-T substitution at position 162 (G-162-->T) in the promoter region of blaTEM and from the structural modifications resulting from the Met-69-->Leu and Arg-275-->Gln substitutions that characterize IRT-14 beta-lactamase.

Anti-Bacterial Agents↗
PubMed2003

Stochastic EM for estimating the parameters of a multilevel IRT model.

An item response theory (IRT) model is used as a measurement error model for the dependent variable of a multilevel model. The dependent variable is latent but can be measured indirectly by using tests or questionnaires. The advantage of using latent scores as dependent variables of a multilevel model is that it offers the possibility of modelling response variation and measurement error and separating the influence of item difficulty and ability level. The two-parameter normal ogive model is used for the IRT model. It is shown that the stochastic EM algorithm can be used to estimate the parameters which are close to the maximum likelihood estimates. This algorithm is easily implemented. The estimation procedure will be compared to an implementation of the Gibbs sampler in a Bayesian framework. Examples using real data are given.

Educational Status↗
PubMed1997

[Routine detection of beta-lactamases TEM resistant to inhibitors (IRT) and oxacillinases (OXA) in Escherichia coli].

Comparative study of twenty two strains of Escherichia coli producing a well-characterized penicillinase of the OXA type (oxacillinase) or of the IRT type (TEM resistant to inhibitors) evidenced different criteria leading to the distinction of two groups of strains, according to the type of the harboured enzyme. Applied to eleven clinical strains, these criteria were always concordant with the determination of the isoelectric point, of hybridization and of oligotyping using probes for the classification into OXA or IRT type. We therefore propose the measurement of three inhibition diameters on antibiogram (cefepime, mecillinam and ceftazidime) for the routine distinction of the two enzymes. The interest of the easy characterization of these enzymes is reinforced by our findings that, as for all OXA strains, the inoculum effect which is displayed by some third-generation cephalosporins is very important and should be taken into account in the treatment of severe infections.

Anti-Bacterial Agents↗
PubMed1981

In-vivo estimates for the uptake of caesium-137 by cattle grazing contaminated pasture around the ESK and IRT estuaries, Cumbria, U.K.

Sea water contaminated with diluted radioactive effluent from the Windscale nuclear complex in Cumbria periodically floods low-lying grazing pasture around the estuaries of the rivers Esk, Irt and Mite near Ravenglass. During 1979, an experiment was carried out to measure the transfer of caesium-137 from grass to muscle in cows grazing these pastures. Grass samples were taken in a vivo external gamma-ray measurements were made on cattle. A very low transfer coefficient was found, less than 9 X 10-4 days kg-1 with a best estimate of 4 X 10-4 days kg-1, compared with a more usual value of around 3 X 10-2 days kg-1. The low transfer seems to occur because the bulk of the caesium-137 on the grass is bound to resuspended estuarine surface sediment deposited during flooding. In this form, the caesium-137 is only poorly absorbed across the gut of the grazing cattle.

Animals↗
PubMed2000

Does computerizing paper-and-pencil job attitude scales make a difference? New IRT analyses offer insight.

The measurement equivalence of 2 scales of the Job Descriptive Index (JDI; P. C. Smith, L. M. Kendall, & C. L. Hulin, 1969), the Supervisor Satisfaction scale and the Coworker Satisfaction scale, was examined across computerized and paper-and-pencil administrations. In this study, employees in 2 organizations (N = 1,777) were administered paper-and-pencil versions of the scales, and employees in a third organization (N = 509) were administered a computerized version. A newly developed item response theory (IRT) technique for examining differential test functioning (N. S. Raju, W. J. van der Linden, & P. F. Fleer, 1995) was used to examine measurement equivalence across media. Results support the measurement equivalence of the JDI Supervisor and Coworker scales across administration media. The implications of these findings for both practitioners and organizational researchers are discussed.

Attitude↗
PubMed2003

Differential regulation of nramp and irt metal transporter genes in wild type and iron uptake mutants of tomato.

Metal transporters regulated by iron can transport a variety of divalent metals, suggesting that iron regulation is important for specificity of iron transport. In plants, the iron-regulated broad-range metal transporter IRT1 is required for uptake of iron into the root epidermis. Functions of other iron-regulated plant metal transporters are not yet established. To deduce novel plant iron transport functions we studied the regulation of four tomato metal transporter genes belonging to the nramp and irt families with respect to environmental and genetic factors influencing iron uptake. We isolated Lenramp1 and Lenramp3 from tomato and demonstrate that these genes encode functional NRAMP metal transporters in yeast, where they were iron-regulated and localized mainly to intracellular vesicles. Lenramp1 and Leirt1 revealed both root-specific expression and up-regulation by iron deficiency, respectively, in contrast to Leirt2 and Lenramp3. Lenramp1 and Leirt1, but not Lenramp3 and Leirt2, were down-regulated in the roots of fer mutant plants deficient in a bHLH gene regulating iron uptake. In chloronerva mutant plants lacking the functional enzyme for synthesis of the plant-specific metal chelator nicotianamine Leirt1 and Lenramp1 were up-regulated despite sufficient iron supply independent of a functional fer gene. Lenramp1 was expressed in the vascular root parenchyma in a similar cellular pattern as the fer gene. However, the fer gene was not sufficient for inducing Lenramp1 and Leirt1 when ectopically expressed. Based on our results, we suggest a novel function for NRAMP1 in mobilizing iron in the vascular parenchyma upon iron deficiency in plants. We discuss fer/nicotianamine synthase-dependent and -independent regulatory pathways for metal transporter gene regulation.

Alkyl and Aryl Transferases↗
PubMed1999

Molecular characterization of TEM-59 (IRT-17), a novel inhibitor-resistant TEM-derived beta-lactamase in a clinical isolate of Klebsiella oxytoca.

A clinical isolate of Klebsiella oxytoca (Kox 443) was found to have a low-level resistance to broad-spectrum penicillins (MICs of amoxicillin and ticarcillin, 256 and 32 microg/ml, respectively), without substantial potentiation by 2 microg of clavulanic acid per ml (amoxicillin- and ticarcillin-clavulanate, 128 and 8 microg/ml, respectively), while being fully susceptible to cephalosporins and other beta-lactam antibiotics. These resistances were carried by a ca. 50-kb conjugative plasmid that encodes a single beta-lactamase with a pI of 5.6. Compared to TEM-2, this enzyme exhibited a 3- to 30-fold higher Km and a decreased maximal hydrolysis rate for beta-lactams; higher concentrations of suicide inactivators (5- to 500-fold higher concentrations giving a 50% reduction in hydrolysis) were required for inhibition. Nucleotide sequence analysis revealed identity between the blaTEM gene of Kox 443 and the blaTEM-2 gene, except for a single A-to-G change at position 590, leading to the amino acid change from Ser-130 Gly. This mutation has not been reported previously in the TEM type beta-lactamases produced by clinical strains, and the novel enzyme was called TEM-59 (alternative name IRT-17). This is the first description of an inhibitor-resistant TEM-derived enzyme in the species K. oxytoca.

Aged↗
PubMed2001

TEM-89 beta-lactamase produced by a Proteus mirabilis clinical isolate: new complex mutant (CMT 3) with mutations in both TEM-59 (IRT-17) and TEM-3.

TEM-89 (CMT-3) is the first complex mutant beta-lactamase produced by a clinical strain of Proteus mirabilis (strain Pm 631). This new enzyme, which has a pI of 6.28, is derived from TEM-3 and has a single amino acid substitution also encountered in TEM-59 (inhibitor-resistant TEM beta-lactamase IRT-17): Ser-130 to Gly. TEM-89 hydrolyzed penicillins to the same extent that TEM-3 did but lost almost all hydrolytic activity for cephalosporins and, like TEM-59, was highly resistant to inhibitors.

Amino Acid Substitution↗
PubMed2005

TEM-109 (CMT-5), a natural complex mutant of TEM-1 beta-lactamase combining the amino acid substitutions of TEM-6 and TEM-33 (IRT-5).

Escherichia coli CF349 exhibited a complex beta-lactam resistance phenotype, including resistance to amoxicillin and ticarcillin alone and in combination with clavulanate and to some extended-spectrum cephalosporins. The double-disk synergy test was positive. CF349 harbored an 85-kb conjugative plasmid which encoded a beta-lactamase of pI 5.9. The corresponding bla gene was identified by PCR and sequencing as a bla(TEM) gene. The deduced protein sequence revealed a new complex mutant of TEM-1 beta-lactamase designated TEM-109 (CMT-5). TEM-109 contained both the substitutions Glu104Lys and Arg164His of the expanded-spectrum beta-lactamase (ESBL) TEM-6 and Met69Leu of the inhibitor-resistant TEM-33 (IRT-5). TEM-109 exhibited hydrolytic activity against ceftazidime similar to that of TEM-6 (k(cat), 56 s(-1) and 105 s(-1), respectively; K(m) values, 226 and 247 microM, respectively). The 50% inhibitory concentrations of clavulanate and tazobactam (0.13 microM and 0.27 microM, respectively) were 5- to 10-fold higher for TEM-109 than for TEM-6 (0.01 and 0.06 microM, respectively) but were almost 10-fold lower than those for TEM-33. The characterization of this novel CMT, which exhibits a low level of resistance to inhibitors, highlights the emergence of this new ESBL type.

Amino Acid Substitution↗