Comparison of Volumetric Analysis Methods for Machine Tools with Rotary Axes

Confidence in the ability of a production machine to meet manufacturing tolerances requires a full understanding of the accuracy of the machine. However, the definition of “the accuracy of the machine” is open to interpretation. Historically, this has been in terms of linear positioning accuracy of an axis with no regard for the other errors of the machine. Industry awareness of the three-dimensional positioning accuracy of a machine over its working envelope has slowly developed to an extent that people are aware that “volumetric accuracy” gives a better estimation of machine performance. However, at present there is no common standard for volumetric errors of machine tools, although several researchers have developed models to predict the effect of the combined errors. The error model for machines with three Cartesian axes has been well addressed, for example by the use of homogenous transformation matrices. Intuitively, the number of error sources increases with the number of axes present on the machine. The effect of the individual axis geometric errors can become increasingly significant as the chain of dependent axes is extended. Measurement of the “volumetric error” or its constituents is often restricted to a subset of the errors of the Cartesian axes by solely relying on a laser interferometer for measurement. This leads to a volumetric accuracy figure that neglects the misalignment errors of rotary axes. In more advanced models the accuracy of the rotary axes are considered as a separate geometric problem whose volumetric accuracy is then added to the volumetric accuracy of the Cartesian axes. This paper considers the geometric errors of some typical machine configurations with both Cartesian and non-Cartesian axes and uses case studies to emphasise the importance of measurement of all the error constituents. Furthermore, it shows the misrepresentation when modelling a five-axis machine as a three-plus-two error problem. A method by which the five-axis model can be analysed to better represent the machine performance is introduced. Consideration is also given for thermal and non-rigid influences on the machine volumetric accuracy analysis, both in terms of the uncertainty of the model and the uncertainty during the measurement. The magnitude of these errors can be unexpectedly high and needs to be carefully considered whenever testing volumetric accuracy, with additional tests being recommended

Frequencies of Lipopolysaccharide Core Types among Clinical Isolates of Escherichia coli Defined with Monoclonal Antibodies

Mouse monoclonal antibodies (MAbs) specific for the lipopolysaccharide (LPS) core types R1, R2, and R3 of Escherichia coli and a cross-reactive MAb that binds to the LPS core of almost all E. coli were used in ELISA to determine the frequency of cores resembling R1, R2, and R3 in strains of E. coli isolated from clinical samples (blood and urine specimens) and from the feces of asymptomatic individuals. Of the 180 wild-type isolates, 123 were assigned to R1 core type, 14 to R2, and 18 to R3. Twenty-five wild-type E. coli isolates could not be assigned to a particular core type and may have either an R4 or K12 core or a previously unrecognized core type. R1 core type was associated with O types 1, 4, 6, 8, and 18 and with K1 or K5 capsules. R3 was associated with O15.O75 isolates could be of either R1 or R2 core typ

Bezlotoxumab for Prevention of Recurrent Clostridium difficile Infection

BACKGROUND Clostridium difficile is the most common cause of infectious diarrhea in hospitalized patients. Recurrences are common after antibiotic therapy. Actoxumab and bezlotoxumab are human monoclonal antibodies against C. difficile toxins A and B, respectively. METHODS We conducted two double-blind, randomized, placebo-controlled, phase 3 trials, MODIFY I and MODIFY II, involving 2655 adults receiving oral standard-of-care antibiotics for primary or recurrent C. difficile infection. Participants received an infusion of bezlotoxumab (10 mg per kilogram of body weight), actoxumab plus bezlotoxumab (10 mg per kilogram each), or placebo; actoxumab alone (10 mg per kilogram) was given in MODIFY I but discontinued after a planned interim analysis. The primary end point was recurrent infection (new episode after initial clinical cure) within 12 weeks after infusion in the modified intention-to-treat population. RESULTS In both trials, the rate of recurrent C. difficile infection was significantly lower with bezlotoxumab alone than with placebo (MODIFY I: 17% [67 of 386] vs. 28% [109 of 395]; adjusted difference, −10.1 percentage points; 95% confidence interval [CI], −15.9 to −4.3; P<0.001; MODIFY II: 16% [62 of 395] vs. 26% [97 of 378]; adjusted difference, −9.9 percentage points; 95% CI, −15.5 to −4.3; P<0.001) and was significantly lower with actoxumab plus bezlotoxumab than with placebo (MODIFY I: 16% [61 of 383] vs. 28% [109 of 395]; adjusted difference, −11.6 percentage points; 95% CI, −17.4 to −5.9; P<0.001; MODIFY II: 15% [58 of 390] vs. 26% [97 of 378]; adjusted difference, −10.7 percentage points; 95% CI, −16.4 to −5.1; P<0.001). In prespecified subgroup analyses (combined data set), rates of recurrent infection were lower in both groups that received bezlotoxumab than in the placebo group in subpopulations at high risk for recurrent infection or for an adverse outcome. The rates of initial clinical cure were 80% with bezlotoxumab alone, 73% with actoxumab plus bezlotoxumab, and 80% with placebo; the rates of sustained cure (initial clinical cure without recurrent infection in 12 weeks) were 64%, 58%, and 54%, respectively. The rates of adverse events were similar among these groups; the most common events were diarrhea and nausea. CONCLUSIONS Among participants receiving antibiotic treatment for primary or recurrent C. difficile infection, bezlotoxumab was associated with a substantially lower rate of recurrent infection than placebo and had a safety profile similar to that of placebo. The addition of actoxumab did not improve efficacy. (Funded by Merck; MODIFY I and MODIFY II ClinicalTrials.gov numbers, NCT01241552 and NCT01513239.

Comparative genomics of European Avian Pathogenic E. coli (APEC)

Background Avian pathogenic Escherichia coli (APEC) causes colibacillosis, which results in significant economic losses to the poultry industry worldwide. However, the diversity between isolates remains poorly understood. Here, a total of 272 APEC isolates collected from the United Kingdom (UK), Italy and Germany were characterised using multiplex polymerase chain reactions (PCRs) targeting 22 equally weighted factors covering virulence genes, R-type and phylogroup. Following these analysis, 95 of the selected strains were further analysed using Whole Genome Sequencing (WGS). Results The most prevalent phylogroups were B2 (47%) and A1 (22%), although there were national differences with Germany presenting group B2 (35.3%), Italy presenting group A1 (53.3%) and UK presenting group B2 (56.1%) as the most prevalent. R-type R1 was the most frequent type (55%) among APEC, but multiple R-types were also frequent (26.8%). Following compilation of all the PCR data which covered a total of 15 virulence genes, it was possible to build a similarity tree using each PCR result unweighted to produce 9 distinct groups. The average number of virulence genes was 6-8 per isolate, but no positive association was found between phylogroup and number or type of virulence genes. A total of 95 isolates representing each of these 9 groupings were genome sequenced and analysed for in silico serotype, Multilocus Sequence Typing (MLST), and antimicrobial resistance (AMR). The UK isolates showed the greatest variability in terms of serotype and MLST compared with German and Italian isolates, whereas the lowest prevalence of AMR was found for German isolates. Similarity trees were compiled using sequencing data and notably single nucleotide polymorphism data generated ten distinct geno-groups. The frequency of geno-groups across Europe comprised 26.3% belonging to Group 8 representing serogroups O2, O4, O18 and MLST types ST95, ST140, ST141, ST428, ST1618 and others, 18.9% belonging to Group 1 (serogroups O78 and MLST types ST23, ST2230), 15.8% belonging to Group 10 (serogroups O8, O45, O91, O125ab and variable MLST types), 14.7% belonging to Group 7 (serogroups O4, O24, O35, O53, O161 and MLST type ST117) and 13.7% belonging to Group 9 (serogroups O1, O16, O181 and others and MLST types ST10, ST48 and others). The other groups (2, 3, 4, 5 and 6) each contained relatively few strains. However, for some of the genogroups (e.g. groups 6 and 7) partial overlap with SNPs grouping and PCR grouping (matching PCR groups 8 (13 isolates on 22) and 1 (14 isolates on 16) were observable). However, it was not possible to obtain a clear correlation between genogroups and unweighted PCR groupings. This may be due to the genome plasticity of E. coli that enables strains to carry the same virulence factors even if the overall genotype is substantially different. Conclusions The conclusion to be drawn from the lack of correlations is that firstly, APEC are very diverse and secondly, it is not possible to rely on any one or more basic molecular or phenotypic tests to define APEC with clarity, reaffirming the need for whole genome analysis approaches which we describe here. This study highlights the presence of previously unreported serotypes and MLSTs for APEC in Europe. Moreover, it is a first step on a cautious reconsideration of the merits of classical identification criteria such as R typing, phylogrouping and serotyping

Generation of Markerless Deletions in the Nosocomial Pathogen <em style="font-family: inherit; font-size: inherit; -webkit-font-smoothing: antialiased; margin: 0px; padding: 0px; border: 0px; outline: 0px; vertical-align: baseline; font-variant: inherit; font-weight: inherit; font-stretch: inherit; line-height: inherit;">Clostridium </em><em style="font-family: inherit; font-size: inherit; letter-spacing: -0.03em; -webkit-font-smoothing: antialiased; margin: 0px; padding: 0px; border: 0px; outline: 0px; vertical-align: baseline; font-variant: inherit; font-weight: inherit; font-stretch: inherit; line-height: inherit;">difficile</em> by induction of DNA Double-strand Breaks

Clostridium difficile is an important nosocomial pathogen associated with potentially fatal disease induced by the use of antibiotics. Genetic characterisation of such clinically important bacteria is often hampered by availability of suitable tools. Here we describe the use of I-SceI to induce DNA double-strand breaks which increase the frequency of allelic exchange and enable the generation of markerless deletions in C. difficile. The usefulness of the system is illustrated by deletion of genes encoding putative AddAB homologues. The ΔaddAB mutants are sensitive to ultra violet light and the antibiotic metronidazole, indicating a role in homologous recombination and repair of DNA breaks. Despite the impairment in recombination, the mutants are still proficient for induction of the SOS response. In addition, deletion of the fliC gene, and subsequent complementation, reveals the importance of potential regulatory elements required for expression of a downstream gene encoding the flagellin glycosyl transferase.IMPORTANCE Most sequenced bacterial genomes contain genes encoding proteins of unknown or hypothetical function. To identify a phenotype for mutations in such genes, deletion is the preferred method for mutagenesis because it reduces the likelyhood of polar effects, although does not eliminate the possibility. Allelic exchange to produce deletions is dependent on the length of homology used to generate merodiploids. Shorter regions of homology resolve at lower frequencies. The work presented here demonstrates the utility of inducing DNA double-strand breaks to increase the frequency of merodiploid resolution in Clostridium difficile Using this approach we reveal the role of two genes, encoding homologues of AddAB, in survival following DNA damage. The method is readily applicable to production of deletions in C. difficile and expands the toolbox available for genetic analysis of this important anaerobic pathogen.</p