Prediction of complete remission and survival in acute myeloid leukemia using supervised machine learning.

Achievement of complete remission signifies a crucial milestone in the therapy of acute myeloid leukemia (AML) while refractory disease is associated with dismal outcomes. Hence, accurately identifying patients at risk is essential to tailor treatment concepts individually to disease biology. We used nine machine learning (ML) models to predict complete remission and 2-year overall survival in a large multicenter cohort of 1,383 AML patients who received intensive induction therapy. Clinical, laboratory, cytogenetic and molecular genetic data were incorporated and our results were validated on an external multicenter cohort. Our ML models autonomously selected predictive features including established markers of favorable or adverse risk as well as identifying markers of so-far controversial relevance. De novo AML, extramedullary AML, double-mutated CEBPA, mutations of CEBPA-bZIP, NPM1, FLT3-ITD, ASXL1, RUNX1, SF3B1, IKZF1, TP53, and U2AF1, t(8;21), inv(16)/t(16;16), del(5)/del(5q), del(17)/del(17p), normal or complex karyotypes, age and hemoglobin concentration at initial diagnosis were statistically significant markers predictive of complete remission, while t(8;21), del(5)/del(5q), inv(16)/t(16;16), del(17)/del(17p), double-mutated CEBPA, CEBPA-bZIP, NPM1, FLT3-ITD, DNMT3A, SF3B1, U2AF1, and TP53 mutations, age, white blood cell count, peripheral blast count, serum lactate dehydrogenase level and hemoglobin concentration at initial diagnosis as well as extramedullary manifestations were predictive for 2-year overall survival. For prediction of complete remission and 2-year overall survival areas under the receiver operating characteristic curves ranged between 0.77-0.86 and between 0.63-0.74, respectively in our test set, and between 0.71-0.80 and 0.65-0.75 in the external validation cohort. We demonstrated the feasibility of ML for risk stratification in AML as a model disease for hematologic neoplasms, using a scalable and reusable ML framework. Our study illustrates the clinical applicability of ML as a decision support system in hematology.

Synthesis, biological evaluation, and molecular docking studies of aldotetronic acid-based LpxC inhibitors.

In order to develop novel inhibitors of the bacterial deacetylase LpxC bearing a substituent to target the UDP binding site of the enzyme, a series of aldotetronic acid-based hydroxamic acids was accessed in chiral pool syntheses starting from 4,6-O-benzylidene-d-glucose and l-arabinitol. The synthesized hydroxamic acids were tested for LpxC inhibitory activity in vitro, revealing benzyl ether 17a ((2S,3S)-4-(benzyloxy)-N,3-dihydroxy-2-[(4-{[4-(morpholinomethyl)phenyl]ethynyl}benzyl)oxy]butanamide) as the most potent LpxC inhibitor. This compound was additionally tested for antibacterial activity against a panel of clinically relevant Gram-negative bacteria, bacterial uptake, and susceptibility to efflux pumps. Molecular docking studies were performed to rationalize the observed structure-activity relationships.

Antibacterial activity of xylose-derived LpxC inhibitors - Synthesis, biological evaluation and molecular docking studies.

LpxC inhibitors represent a promising class of novel antibiotics selectively combating Gram-negative bacteria. In chiral pool syntheses starting from D- and L-xylose, a series of four 2r,3c,4t-configured C-furanosidic LpxC inhibitors was obtained. The synthesized hydroxamic acids were tested for antibacterial and LpxC inhibitory activity, the acquired biological data were compared with those of previously synthesized C-furanosides, and molecular docking studies were performed to rationalize the observed structure-activity relationships. Additionally, bacterial uptake and susceptibility to efflux pump systems were investigated for the most promising stereoisomers.

Detection of a Novel gyrB Mutation Associated With Fluoroquinolone-Nonsusceptible Salmonella enterica serovar Typhimurium Isolated From a Bloodstream Infection in Ghana.

A multidrug-resistant Salmonella enterica serovar Typhimurium with reduced susceptibility to ciprofloxacin was isolated from the blood of a hospitalized child in Ghana. DNA sequencing identified a novel gyrB mutation at codon 466 (Glu466Asp). An increase in fluoroquinolone susceptibility after the introduction of a wild-type gyrB(+) allele demonstrated that the gyrB466 mutation had a direct effect on fluoroquinolone susceptibility.

The Emergence of Reduced Ciprofloxacin Susceptibility in Salmonella enterica Causing Bloodstream Infections in Rural Ghana.

BACKGROUND: Salmonella ranks among the leading causes of bloodstream infections in sub-Saharan Africa. Multidrug resistant typhoidal and nontyphoidal Salmonella (NTS) isolates have been previously identified in this region. However, resistance to ciprofloxacin has rarely been reported in West Africa. This study aims to assess susceptibility against ciprofloxacin in Salmonella causing invasive bloodstream infections among children in rural Ghana. METHODS: From May 2007 until May 2012, children attending a rural district hospital in central Ghana were eligible for recruitment. Salmonella enterica isolated from blood cultures were assessed for ciprofloxacin susceptibility by Etest (susceptible minimum inhibitory concentration [MIC] ≤ 0.06 µg/mL). The gyrA, gyrB, parC, and parE genes were sequenced to identify mutations associated with changes in susceptibility to fluoroquinolones. RESULTS: Two hundred eighty-five Salmonella enterica isolates from 5211 blood cultures were most commonly identified as Salmonella enterica serovar Typhimurium (n = 129 [45%]), Salmonella enterica serovar Typhi (n = 89 [31%]), Salmonella enterica serovar Dublin (n = 20 [7%]), and Salmonella enterica serovar Enteritidis (n = 19 [7%]). All S. Typhi and S. Dublin were susceptible to ciprofloxacin. Reduced susceptibility (MIC >0.06 µg/mL) was found in 53% (10/19) of S. Enteritidis and in 2% (3/129) of S. Typhimurium isolates. Sequencing detected a single gyrB mutation (Glu466Asp) and a single gyrA mutation (Ser83Tyr) in all 3 S. Typhimurium isolates, while 9 of 10 S. Enteritidis harbored single gyrA mutations (Asp87Gly, Asp87Asn, or Asp87Tyr). No mutations were found in the parC and parE genes. CONCLUSIONS: Ciprofloxacin susceptibility in invasive NTS in rural Ghana is highly dependent on serotype. Although reduced ciprofloxacin susceptibility is low in S. Typhimurium, more than half of all S. Enteritidis isolates are affected. Healthcare practitioners in Ghana should be aware of potential treatment failure in patients with invasive S. Enteritidis infections.

Unsupervised meta-clustering identifies risk clusters in acute myeloid leukemia based on clinical and genetic profiles.

BACKGROUND: Increasingly large and complex biomedical data sets challenge conventional hypothesis-driven analytical approaches, however, data-driven unsupervised learning can detect inherent patterns in such data sets. METHODS: While unsupervised analysis in the medical literature commonly only utilizes a single clustering algorithm for a given data set, we developed a large-scale model with 605 different combinations of target dimensionalities as well as transformation and clustering algorithms and subsequent meta-clustering of individual results. With this model, we investigated a large cohort of 1383 patients from 59 centers in Germany with newly diagnosed acute myeloid leukemia for whom 212 clinical, laboratory, cytogenetic and molecular genetic parameters were available. RESULTS: Unsupervised learning identifies four distinct patient clusters, and statistical analysis shows significant differences in rate of complete remissions, event-free, relapse-free and overall survival between the four clusters. In comparison to the standard-of-care hypothesis-driven European Leukemia Net (ELN2017) risk stratification model, we find all three ELN2017 risk categories being represented in all four clusters in varying proportions indicating unappreciated complexity of AML biology in current established risk stratification models. Further, by using assigned clusters as labels we subsequently train a supervised model to validate cluster assignments on a large external multicenter cohort of 664 intensively treated AML patients. CONCLUSIONS: Dynamic data-driven models are likely more suitable for risk stratification in the context of increasingly complex medical data than rigid hypothesis-driven models to allow for a more personalized treatment allocation and gain novel insights into disease biology.

There are various ways in which clinicians can predict the risk of disease progression in patients with leukemia, helping them to treat the patients accordingly. However, these approaches are usually designed by human experts and might not fully capture the complexity of a patient's disease. Here, with a large cohort of patients with acute myeloid leukemia, we design an unsupervised machine learning model ­ a type of computer model that learns from patterns in data without human input­to separate these patients into subgroups according to risk. We identify four distinct groups which differ with regards to patient genetics, laboratory values, and clinical characteristics. These groups have differences in response to treatment and patient survival, and we validate our findings in another dataset. Our approach might help clinicians to better predict outcomes in patients with leukemia and make decisions on treatment.

Prominence of an O75 clonal group (clonal complex 14) among non-ST131 fluoroquinolone-resistant Escherichia coli causing extraintestinal infections in humans and dogs in Australia.

Fluoroquinolone (FQ)-resistant extraintestinal pathogenic Escherichia coli (FQ(r) ExPEC) strains from phylogenetic group B2 are undergoing epidemic spread. Isolates belonging to phylogenetic group B2 are generally more virulent than other E. coli isolates; therefore, resistance to FQs among group B2 isolates is concerning. Although clonal expansion of sequence type 131 (ST131) is a major factor, the contribution of additional clonal groups has not been quantified. Group B2 FQ(r) ExPEC isolates from humans (n = 250) and dogs (n = 12) in Australia were screened for ST131, a recently recognized and rapidly emerging multidrug-resistant and virulent clonal group that is important in both human and companion animal medicine. Non-ST131 isolates underwent virulence genotyping, PCR-based O typing, partial multilocus sequence typing (MLST), pulsed-field gel electrophoresis (PFGE), and FQ resistance mechanism analysis. Of 49 non-ST131 isolates (45 human, 4 canine), 49% (24 human, 2 canine) represented O-type O75 and exhibited conserved virulence genotypes (F10 papA allele, iha, fimH, sat, vat, fyuA, iutA, kpsMII, usp, ompT, malX, K1/K5 capsule) and MLST allele profiles corresponding with clonal complex CC14. Two clusters, each containing canine and human isolates, were identified by PFGE (differentiated by K1 and K5 capsules). Australian FQ(r) O75 isolates exhibited commonality with an historical FQ-susceptible O75 urosepsis isolate (also CC14). The isolation from humans and dogs of highly similar FQ(r) derivatives of the classic O75:K1/K5 (CC14) ExPEC lineage suggests recent acquisition of FQ resistance and potential cross-host-species transfer. This lineage should be targeted with ST131 in future epidemiological investigations of FQ(r) ExPEC.

Commonality among fluoroquinolone-resistant sequence type ST131 extraintestinal Escherichia coli isolates from humans and companion animals in Australia.

Escherichia coli sequence type 131 (ST131), an emergent multidrug-resistant extraintestinal pathogen, has spread epidemically among humans and was recently isolated from companion animals. To assess for human-companion animal commonality among ST131 isolates, 214 fluoroquinolone-resistant extraintestinal E. coli isolates (205 from humans, 9 from companion animals) from diagnostic laboratories in Australia, provisionally identified as ST131 by PCR, selectively underwent PCR-based O typing and bla(CTX-M-15) detection. A subset then underwent multilocus sequence typing (MLST), pulsed-field gel electrophoresis (PFGE) analysis, extended virulence genotyping, antimicrobial susceptibility testing, and fluoroquinolone resistance genotyping. All isolates were O25b positive, except for two O16 isolates and one O157 isolate, which (along with six O25b-positive isolates) were confirmed by MLST to be ST131. Only 12% of isolates (25 human, 1 canine) exhibited bla(CTX-M-15). PFGE analysis of 20 randomly selected human and all 9 companion animal isolates showed multiple instances of ≥94% profile similarity across host species; 12 isolates (6 human, 6 companion animal) represented pulsotype 968, the most prevalent ST131 pulsotype in North America (representing 23% of a large ST131 reference collection). Virulence gene and antimicrobial resistance profiles differed minimally, without host species specificity. The analyzed ST131 isolates also exhibited a conserved, host species-independent pattern of chromosomal fluoroquinolone resistance mutations. However, eight (89%) companion animal isolates, versus two (10%) human isolates, possessed the plasmid-borne qnrB gene (P < 0.001). This extensive across-species strain commonality, plus the similarities between Australian and non-Australian ST131 isolates, suggest that ST131 isolates are exchanged between humans and companion animals both within Australia and intercontinentally.

Antibacterial activity of finafloxacin under different pH conditions against isogenic strains of Escherichia coli expressing combinations of defined mechanisms of fluoroquinolone resistance.

OBJECTIVES: Finafloxacin is an investigational fluoroquinolone exhibiting broad-spectrum activity that is enhanced under slightly acidic conditions (pH 5.0-6.5). The impact of individual and combinations of chromosomal mutations (gyrA, parC and marR) and the plasmid-mediated fluoroquinolone resistance mechanisms QepA1, QnrA1, QnrB1, QnrS1 and AAC(6')-Ib-cr were investigated. METHODS: The MICs of finafloxacin, compared with those of ciprofloxacin, levofloxacin and moxifloxacin, were determined at pH 5.8 and 7.2. RESULTS: MICs of finafloxacin compared with other fluoroquinolones at pH 5.8 were lower by a factor of 2-256. MICs of finafloxacin were unaffected by QepA1. Moreover, finafloxacin appeared not to be a substrate for AAC(6')-Ib-cr. CONCLUSIONS: Compared with ciprofloxacin, levofloxacin and moxifloxacin, finafloxacin shows higher activity especially at pH 5.8 against Escherichia coli mutants expressing known fluoroquinolone resistance determinants alone and in combinations.

Alcohols for skin antisepsis at clinically relevant skin sites.

The antiseptic efficacy of ethanol, isopropanol, and n-propanol at 60%, 70%, and 89.5% (all vol/vol) was analyzed after 2, 3, or 4 min of application to the forehead, back, and abdomen of 180 volunteers by the use of a standardized swab sampling method. Results of recolonization by the aerobic skin flora of the upper arms and backs of 20 volunteers were compared 72 h after treatment with 0.5%, 1%, or 2% chlorhexidine digluconate (CHG) in 89.5% n-propanol. The most effective alcohol at all skin sites was n-propanol, with a mean log(10) reduction of 1.82 after 2 min on the forehead. Efficacy against the aerobic flora of the forehead was mainly influenced by the type of alcohol (P < 0.001), followed by the concentration (P < 0.001) and the application time (P = 0.006). Ethanol and isopropanol were significantly less effective (both P < 0.001). Alcohol supplemented with 0.5% or more CHG was significantly more effective than alcohol alone in the suppression of recolonization (P < 0.05). An 89.5% solution of n-propanol was the most effective alcohol for the reduction of populations of aerobic skin flora. Its combination with CHG is appropriate whenever recolonization of the skin must be limited. Further studies are needed to determine the most effective concentration of CHG in n-propanol to provide the best protection against recolonization of the skin, e.g., for catheter site care.

The role of intra- and extragenic compensatory mutations in the suppression of fluoroquinolone resistance in a Salmonella Typhimurium gyrA mutant (D87G).

OBJECTIVES: The live vaccine strain TAD Salmonella vacT (vacT) carrying gyrA mutations W59R, G75A, D87G and A866S shows resistance to nalidixic acid and rifampicin, but increased susceptibility to macrolides, fluoroquinolones and phenylalanyl-arginyl-beta-naphthylamide. This phenotype contrasts with the presence of the gyrA mutation D87G usually associated with reduced susceptibility to fluoroquinolones. Thus, a possible compensatory effect on the suppression of gyrA-mediated resistance by gyrA mutations within the quinolone resistance-determining region alone or in combination (intragenic) or by a mutation affecting AcrAB-TolC (extragenic), the major multidrug resistance efflux pump in Salmonella, was investigated. METHODS: Site-specifically introducing combinations of novel mutations into the chromosomal gyrA gene of the vacT parent strain M415 yielded defined gyrA mutants. These were characterized by determining fluoroquinolone susceptibilities and the degree of DNA supercoiling. DNA sequences of acrR, acrA and acrB of vacT were determined. A role of acrB mutations in increased fluoroquinolone susceptibility of vacT was investigated by a complementation test. RESULTS: All in vitro generated mutants carrying the gyrA D87G mutation showed increased fluoroquinolone MICs and reduced degrees of DNA supercoiling, irrespective of the presence of an additional gyrA mutation, W59R or G75A. DNA sequence analysis revealed two deletions in acrB of vacT. Complementation of vacT with an acrB wild-type gene decreased the susceptibilities to fluoroquinolones and macrolides. CONCLUSIONS: The increased fluoroquinolone susceptibility of the gyrA mutant vacT is most plausibly explained by a reduced efflux pump activity caused by acrB deletions in vacT.

Type II topoisomerases--inhibitors, repair mechanisms and mutations.

Type II topoisomerases are ubiquitous enzymes that play an essential role in the control of replicative DNA synthesis and share structural and functional homology among different prokaryotic and eukaryotic organisms. Antibacterial fluoroquinolones target prokaryotic topoisomerases at concentrations 100- to 1000-fold lower than mammalian enzymes, the preferred targets of anticancer drugs such as etoposide. The mechanisms of action of both of these types of inhibitors involve the fixation of an intermediate reaction step, where the enzyme is covalently bound to an enzyme-mediated DNA double-strand break (DSB). The resulting ternary drug-enzyme-DNA complexes can then be converted to cleavage complexes that block further movement of the DNA replication fork, subsequently inducing stress responses. In haploid prokaryotic cells, stress responses include error-free and error-prone DNA damage repair pathways, such as homologous recombination and translesion synthesis, respectively. The latter can result in the acquisition of point mutations. Diploid mammalian cells are assumed to preferentially use recombination mechanisms for the repair of DSBs, an example of which, non-homologous end joining, is a major error-prone repair mechanism associated with an increased frequency of detectable small deletions, insertions and translocations. However, results obtained from safety testing of novel fluoroquinolones at high concentrations indicate that point mutations may also occur in mammalian cells. Recent data provide evidence for translesion synthesis catalysed by error-prone repair polymerases as a damage-tolerance repair mechanism of DSBs in eukaryotic cells. This paper discusses possible roles of different mechanisms for the repair of DSBs operating in both eukaryotic and prokaryotic cells that result in recombinational rearrangements, deletions/insertions as well as point mutations.

Pitfalls in efficacy testing--how important is the validation of neutralization of chlorhexidine digluconate?

BACKGROUND: Effective neutralization of active agents is essential to obtain valid efficacy results, especially when non-volatile active agents like chlorhexidine digluconate (CHG) are tested. The aim of this study was to determine an effective and non-toxic neutralizing mixture for a propan-1-ol solution containing 2% CHG. METHODS: Experiments were carried out according to ASTM E 1054-02. The neutralization capacity was tested separately with five challenge microorganisms in suspension, and with a rayon swab carrier. Either 0.5 mL of the antiseptic solution (suspension test) or a saturated swab with the antiseptic solution (carrier test) was added to tryptic soy broth containing neutralizing agents. After the samples were mixed, aliquots were spread immediately and after 3 h of storage at 2 - 8 degrees C onto tryptic soy agar containing a neutralizing mixture. RESULTS: The neutralizer was, however, not consistently effective in the suspension test. Immediate spread yielded a valid neutralization with Staphylococcus aureus, Staphylococcus epidermidis and Corynebacterium jeikeium but not with Micrococcus luteus (p < 0.001) and Candida albicans (p < 0.001). A 3-h storage period of the neutralized active agents in suspension resulted in significant carry-over activity of CHG in addition against Staphylococcus epidermidis (p < 0.001) and Corynebacterium jeikeium (p = 0.044). In the carrier test, the neutralizing mixture was found to be effective and non toxic to all challenge microorganisms when spread immediately. However, after 3 h storage of the neutralized active agents significant carry-over activity of CHG against Micrococcus luteus (p = 0.004; Tukey HSD) was observed. CONCLUSION: Without effective neutralization in the sampling fluid, non-volatile active ingredients will continue to reduce the number of surviving microorganisms after antiseptic treatment even if the sampling fluid is kept cold straight after testing. This can result in false-positive antiseptic efficacy data. Attention should be paid during the neutralization validation process to the amount of antiseptic solution, the storage time and to the choice of appropriate and sensitive microorganisms.

Investigation of Antifouling Properties of Surfaces Featuring Zwitterionic α-Aminophosphonic Acid Moieties.

Zwitterionic thin films containing α-amino phosphonic acid moieties were successfully introduced on silicon surfaces and their antifouling properties were investigated. Initially, the substrates were modified with a hybrid polymer, composed of poly(methylsilsesquioxane) (PMSSQ) and poly(4-vinyl benzaldehyde) (PStCHO). Next, a Kabachnik-Fields post-polymerization modification (sur-KF-PMR) of the functionalized aldehyde surfaces was conducted with different amines and dialkyl phosphonates. After subsequent deprotection reaction of dialkyl phosphonates, the obtained zwitterionic surfaces were characterized by various techniques and we found excellent antifouling properties of the resulting films.

Topoisomerase IV mutations in quinolone-resistant salmonellae selected in vitro.

The development of high-level fluoroquinolone resistance has rarely been observed in salmonellae and, in contrast to other Gram-negative bacteria mutations affecting topoisomerase IV, a known secondary target of quinolones in Escherichia coli has not been described except for one recent report. The present study used quinolone-susceptible field isolates representing epidemiologically relevant serovars and phage types Salmonella Hadar and Salmonella Typhimurium DT104 and DT204c to select fluoroquinolone-resistant mutants in vitro. Three selection steps were necessary to obtain high-level fluoroquinolone-resistant mutants (MICCip > or = 8 microg/ml). All first-step mutants examined had a single gyrA mutation (affecting either Ser83 or Asp87). Additional topoisomerase mutations affecting gyrA (Asp87), gyrB (Ser464), and parC (Gly78) were detected in second- and third-step mutants. Introducing into the respective mutants the corresponding plasmid-coded quinolone-susceptible allele of either gyrA, gyrB, or parC resulted in reduction of quinolone resistance, indicating a role for these mutations in quinolone resistance. In the presence of an inhibitor of RND-type efflux pumps, the susceptibilities to ciprofloxacin and chloramphenicol of second- and third-step mutants increased by two to four serial dilution steps, providing evidence that an efflux-mediated resistance mechanism contributes to the development of high-level fluoroquinolone resistance in salmonellae.

Beta-lactamase expression in Yersinia enterocolitica biovars 1A, 1B, and 3.

Characteristic patterns of beta-lactam susceptibility are associated with different biovars of Yersinia enterocolitica. In a previous study differences in beta-lactam susceptibility among biovar 2, 4 and 5 strains were largely attributed to differences in expression of beta-lactamase A (BlaA) and beta-lactamase B (BlaB). The basis for differences in beta-lactam susceptibility of strains of biovars 1A, 1B and 3 is now considered. All the strains examined had blaB; nine of 31 biovar 3 strains and two of 13 biovar 1B strains had blaA, but PCR did not amplify blaA from biovar 1A strains. Nevertheless, inhibition data indicated that the majority of uninduced biovar 1A strains expressed BlaA and BlaB in similar amounts. Strong inducibility was seen in all these strains. Biovar 1B strains (which were less inducible than strains of biovar 1A) predominantly produced BlaA without induction; ticarcillin-sensitive strains of biovar 3 produced only BlaB but were not inducible; without induction biovar 3 strains resistant to ticarcillin and amoxycillin/clavulanate produced either predominantly BlaA, predominantly BlaB or exclusively BlaB and induction was demonstrated except for strains producing BlaB alone; biovar 3 strains resistant to ticarcillin but sensitive to amoxycillin/clavulanate predominantly produced BlaA without induction and were inducible for beta-lactamase activity. After induction, nearly all strains predominantly or exclusively produced BlaB. Although PCR amplification fragments with primers specific for blaA were obtained only from some strains, the induction and inhibition data suggest that all Y. enterocolitica strains possess enzymes related to BlaA- as well as BlaB. Nevertheless, expression of the beta-lactamase is regulated differently in different biovars and varies within most biovars. Failure to predict beta-lactamase expression profiles from MIC data indicates the presence of additional mechanisms contributing to differences in susceptibility.

Expression of beta-lactamases in Yersinia enterocolitica strains of biovars 2, 4 and 5.

Characteristic patterns of susceptibility to beta-lactam antibiotics are associated with different biovars of Yersinia enterocolitica. To elucidate the basis for these differences, the beta-lactamases of strains of Y. enterocolitica biovars 4 (n = 63), 2 (n = 12) and 5 (n = 10) were characterised. PCR fragments were generated from the beta-lactamase A (blaA) and B (blaB) genes; in addition, beta-lactamase induction tests were performed with imipenem as the inducer and beta-lactamase inhibition assays were undertaken with aztreonam and clavulanic acid. All the strains yielded PCR amplification fragments with primers to blaA and blaB. Biovar 4 strains had uniform patterns of beta-lactamase induction and inhibition: uninduced biovar 4 strains predominantly expressed BlaA, but low-level expression of BlaB was also detected; after induction, biovar 4 strains predominantly produced BlaB. Beta-lactamase expression varied between and within biovars 2 and 5: uninduced strains predominantly expressed either BlaA or BlaB, or exclusively BlaB; after induction BlaB was predominantly or exclusively expressed. Both the basal and induced levels of beta-lactamase varied within biovars 2 and 5. Some biovar 5 strains were not inducible; these predominantly produced BlaA. The results of this study show that biovar 2, 4 and 5 strains contain both blaA and blaB, but that the expression of the enzymes is regulated differently between the biovars, and varies within biovars 2 and 5. There was some correlation between antibiogram and the clusters defined from the beta-lactamase induction and inhibition tests, but it was not possible to predict beta-lactamase expression profiles from MIC data.

Uncomplicated Urinary Tract Infections and Antibiotic Resistance-Epidemiological and Mechanistic Aspects.

Uncomplicated urinary tract infections are typically monobacterial and are predominantly caused by Escherichia coli. Although several effective treatment options are available, the rates of antibiotic resistance in urinary isolates of E. coli have increased during the last decade. Knowledge of the actual local rates of antibiotic resistant pathogens as well as the underlying mechanisms are important factors in addition to the geographical location and the health state of the patient for choosing the most effective antibiotic treatment. Recommended treatment options include trimethoprim alone or in combination with sulfamethoxazol, fluoroquinolones, ß-lactams, fosfomycin-trometamol, and nitrofurantoin. Three basic mechanisms of resistance to all antibiotics are known, i.e., target alteration, reduced drug concentration and inactivation of the drug. These mechanisms-alone or in combination-contribute to resistance against the different antibiotic classes. With increasing prevalence, combinations of resistance mechanisms leading to multiple drug resistant (mdr) pathogens are being detected and have been associated with reduced fitness under in vitro situations. However, mdr clones among clinical isolates such as E. coli sequence type 131 (ST131) have successfully adapted in fitness and growth rate and are rapidly spreading as a worldwide predominating clone of extraintestinal pathogenic E. coli.

Biomimetic PEG-catecholates for stabile antifouling coatings on metal surfaces: applications on TiO2 and stainless steel.

Trimeric catecholates have been designed for the stable immobilization of effector molecules on metal surfaces. The design of these catecholates followed a biomimetic approach and was inspired by natural multivalent metal binders, such as mussel adhesion proteins (MAPs) and siderophores. Three catecholates have been conjugated to central scaffolds based on adamantyl or trisalkylmethyl core structures. The resulting triscatecholates have been immobilized on TiO2 and stainless steel. In a proof of concept study we have demonstrated the high stability of the resulting nanolayers at neutral and slightly acidic pH. Furthermore, polyethylene glycol (PEG) conjugates of our triscatecholates have been synthesized and were immobilized on TiO2 and stainless steel. The PEG coated surfaces showed excellent antifouling properties upon exposure to human blood and bacteria as demonstrated by fluorescence microscopy, ellipsometry and a bacterial assay with Staphylococcus epidermidis. In addition, our PEG-triscatecholates showed no cytotoxicity against bone-marrow stem cells on TiO2.

CMY-42, a novel plasmid-mediated CMY-2 variant AmpC beta-lactamase.

We isolated a clinical Escherichia coli strain with an antimicrobial resistance phenotype characteristic for the expression of an AmpC beta-lactamase. Molecular methods revealed a novel, plasmid-localized variant of CMY-2 with a substitution of valine 231 for serine (V231S), which was designated CMY-42. Like the CMY-2-like AmpC beta-lactamase CMY-30, carrying the substitution V231G, CMY-42 displayed increased activity toward expanded spectrum cephalosporins. This finding supports the hypothesis that a bulky side chain at position 231 (Ambler's position 211) may pose a steric clash with certain cephalosporins hindering the access of the AmpC beta-lactamase; however, additional phenomena may account for the observed hydrolytic properties.