Discrimination of Aspergillus lentulus from Aspergillus fumigatus by Raman spectroscopy and MALDI-TOF MS.

In 2005, a new sibling species of Aspergillus fumigatus was discovered: Aspergillus lentulus. Both species can cause invasive fungal disease in immune-compromised patients. The species are morphologically very similar. Current techniques for identification are PCR-based or morphology-based. These techniques are labour-intense and not sufficiently discriminatory. Since A. lentulus is less susceptible to several antifungal agents, it is important to correctly identify the causative infectious agent in order to optimize antifungal therapy. In this study we determined whether Raman spectroscopy and/or MALDI-TOF MS were able to differentiate between A. lentulus and A. fumigatus. For 16 isolates of A. lentulus and 16 isolates of A. fumigatus, Raman spectra and peptide profiles were obtained using the Spectracell and MALDI-TOF MS (VITEK MS RUO, bioMérieux) respectively. In order to obtain reliable Raman spectra for A. fumigatus and A. lentulus, the culture medium needed to be adjusted to obtain colourless conidia. Only Raman spectra obtained from colourless conidia were reproducible and correctly identified 25 out of 32 (78 %) of the Aspergillus strains. For VITEK MS RUO, no medium adjustments were necessary. Pigmented conidia resulted in reproducible peptide profiles as well in this case. VITEK MS RUO correctly identified 100 % of the Aspergillus isolates, within a timeframe of approximately 54 h including culture. Of the two techniques studied here, VITEK MS RUO was superior to Raman spectroscopy in the discrimination of A. lentulus from A. fumigatus. VITEK MS RUO seems to be a successful technique in the daily identification of Aspergillus spp. within a limited timeframe.

Prevalence of nasal methicillin-resistant Staphylococcus aureus colonization in healthcare workers in a Western Australian acute care hospital.

Due to a longstanding comprehensive "search and destroy policy", methicillin-resistant Staphylococcus aureus (MRSA) is not endemic in Western Australian (WA) acute care hospitals. As the prevalence of MRSA in the community has increased, healthcare workers (HCW) are at risk of importing MRSA into hospitals. We aimed to determine the prevalence of and risk factors for nasal MRSA colonization in our HCW population. A period prevalence study was conducted at an 850-bed tertiary hospital. Basic demographics and a nasal swab were obtained. A total of 1,542 HCWs employed in our centre were screened for MRSA, of whom 3.4% (n = 52) were colonized. MRSA colonization was more common in patient care assistants (6.8%) and nurses (5.2%) than in allied health professionals (1.7%) and doctors (0.7%) (p < 0.01). Working in "high-risk" wards that cared for MRSA colonized/infected patients was the strongest risk factor for HCW MRSA colonization (p < 0.001). ST1-IV and ST78-IV (the most common community clones in the region) were the most frequently identified clones. In conclusion, MRSA colonization of HCWs occurs primarily in HCWs caring for patients colonized or infected with MRSA. Surveillance screening of HCWs should be regularly performed on wards with patients with high MRSA colonization prevalence to prevent further spread in the hospital.

Experimental and computational morphology of three polymorphs of the free base of Venlafaxine: a comparison of morphology prediction methods.

In this paper the experimental and the computational studies of the morphology of three polymorphs of the free base of Venlafaxine ((N,N-dimethyl)-2-(1-hydroxy cyclohex-1-yl)-2-(4-methoxyphenyl) ethylamine) are reported. The morphology of all polymorphs has been predicted using the Bravais-Friedel-Donnay-Harker method, the attachment energy method and kinetic Monte Carlo growth simulations and these predictions have been compared with experimental observations. The Monte Carlo simulations allow for a detailed simulation of the growth process, including driving force and growth mechanism, which leads to a semi-quantitative prediction of the growth morphologies of all three phases. For phase I two distinct growth habits are found experimentally under the same conditions. This is explained by the occurrence of a spiral growth mechanism in one of the two, which was observed using AFM and which is also supported by the Monte Carlo simulations. The habit of phase II could only be explained from simulations when a spiral growth mechanism is assumed; the shape of phase III could not be modeled accurately from the Monte Carlo simulations. Although the shape of the crystal is reproduced accurately, some of the indices of the faces predicted are not in agreement with the indices measured. The deviations are interpreted to be due to the presence of domains in the crystals as a result of the layered structure.

Crystal structure prediction of organic pigments: quinacridone as an example.

The structures of the alpha, beta and gamma polymorphs of quinacridone (Pigment Violet 19) were predicted using Polymorph Predictor software in combination with X-ray powder diffraction patterns of limited quality. After generation and energy minimization of the possible structures, their powder patterns were compared with the experimental ones. On this basis, candidate structures for the polymorphs were chosen from the list of all structures. Rietveld refinement was used to validate the choice of structures. The predicted structure of the gamma polymorph is in accordance with the experimental structure published previously. Three possible structures for the beta polymorph are proposed on the basis of X-ray powder patterns comparison. It is shown that the alpha structure in the Cambridge Structural Database is likely to be in error, and a new alpha structure is proposed. The present work demonstrates a method to obtain crystal structures of industrially important pigments when only a low-quality X-ray powder diffraction pattern is available.

Epitaxial 2D nucleation of the stable polymorphic form of the steroid 7alphaMNa on the metastable form: implications for Ostwald's rule of stages.

This paper presents in situ observations of the epitaxial nucleation and growth of the stable polymorph of a steroid, 7alphaMna, on a specific face of the metastable form at low supersaturation, using optical microscopy and in situ Raman spectroscopy. The presence of the metastable polymorph is essential for the nucleation and growth of the stable one. The order of the metastable zones of the stable and metastable polymorphs is reversed for the epitaxial growth process as compared to the case of 3D nucleation. The rate of transformation of the metastable polymorph to the stable one can be controlled by the supersaturation.

A third blind test of crystal structure prediction.

Following the interest generated by two previous blind tests of crystal structure prediction (CSP1999 and CSP2001), a third such collaborative project (CSP2004) was hosted by the Cambridge Crystallographic Data Centre. A range of methodologies used in searching for and ranking the likelihood of predicted crystal structures is represented amongst the 18 participating research groups, although most are based on the global minimization of the lattice energy. Initially the participants were given molecular diagrams of three molecules and asked to submit three predictions for the most likely crystal structure of each. Unlike earlier blind tests, no restriction was placed on the possible space group of the target crystal structures. Furthermore, Z' = 2 structures were allowed. Part-way through the test, a partial structure report was discovered for one of the molecules, which could no longer be considered a blind test. Hence, a second molecule from the same category (small, rigid with common atom types) was offered to the participants as a replacement. Success rates within the three submitted predictions were lower than in the previous tests - there was only one successful prediction for any of the three ;blind' molecules. For the ;simplest' rigid molecule, this lack of success is partly due to the observed structure crystallizing with two molecules in the asymmetric unit. As in the 2001 blind test, there was no success in predicting the structure of the flexible molecule. The results highlight the necessity for better energy models, capable of simultaneously describing conformational and packing energies with high accuracy. There is also a need for improvements in search procedures for crystals with more than one independent molecule, as well as for molecules with conformational flexibility. These are necessary requirements for the prediction of possible thermodynamically favoured polymorphs. Which of these are actually realised is also influenced by as yet insufficiently understood processes of nucleation and crystal growth.

Method for the computational comparison of crystal structures.

A new method for assessing the similarity of crystal structures is described. A similarity measure is important in classification and clustering problems in which the crystal structures are the source of information. Classification is particularly important for the understanding of properties of crystals, while clustering can be used as a data reduction step in polymorph prediction. The method described uses a radial distribution function that combines atomic coordinates with partial atomic charges. The descriptor is validated using experimental data from a classification study of clathrate structures of cephalosporins and data from a polymorph prediction run. In both cases, excellent results were obtained.

A test of crystal structure prediction of small organic molecules.

A collaborative workshop was held in May 1999 at the Cambridge Crystallographic Data Centre to test how well currently available methods of crystal structure prediction perform when given only the atomic connectivity for an organic compound. A blind test was conducted on a selection of four compounds and a wide range of methodologies representing, the principal computer programs currently available were used. There were 11 participants who were allowed to propose at most three structures for each compound. No program gave consistently reliable results. However, seven proposed structures were close to an experimental one and were classified as "correct". One compound occurred in two polymorphs, but only one form was predicted correctly among the calculated structures. The basic problem with lattice energy based methods of crystal structure prediction is that many structures are found within a few kJ mol(-1) of the global minimum. The fine detail of the force-field methodology and parametrization influences the energy ranking within each method. Nevertheless, present methods may be useful in providing a set of structures as possible polymorphs for a given molecular structure.

Comparative spectra analysis (CoSA): spectra as three-dimensional molecular descriptors for the prediction of biological activities.

A novel 3D QSAR approach, comparative spectra analysis (CoSA), in which molecular spectra are used as three-dimensional molecular descriptors for the prediction of biological activities, is presented and discussed. To this purpose, experimentally determined 1H NMR, mass, and IR spectra, as well as simulated IR and 13C NMR spectra, for a set of 45 diverse progestagens are converted by a program, SpecMat, into matrixes, which are subsequently employed in a multivariate regression analysis (PLS). The results are compared with those resulting from a comparative molecular field analysis (CoMFA). When used individually, spectral descriptors yield better correlations and predictions than molecular field descriptors. A combination of spectral descriptors with other descriptors, either spectral or molecular field in nature, leads in most cases to models that are statistically superior to the ones obtained by their corresponding individual spectral or molecular field descriptors.