Invasive pneumococcal disease following adult allogeneic hematopoietic stem cell transplantation.

BACKGROUND: Allogeneic hematopoietic stem cell transplantation (alloHSCT) recipients are at high risk of invasive pneumococcal disease (IPD). We investigated the incidence and risk factors of IPD in alloHSCT recipients from 4 regional transplant centers over an 11-year period. This study aimed to inform future improvements in post-transplant care. METHODS: We conducted a retrospective nested 1:2 case-control study in patients aged ≥18 years who underwent alloHSCT between 2001 and 2011 in 4 major allogeneic transplant centers. Controls were matched with IPD cases on the basis of conditioning intensity and donor relationship (related or unrelated). Demographics and clinical characteristics of cases and controls were summarized. Univariate analysis of risk factors in matched case-control sets, and multivariate conditional logistic regression to control for confounding, were performed. RESULTS: In 23 alloHSCT recipients, 26 IPD episodes were identified. The cumulative incidence over 11 years was 2.3% (95% confidence interval [CI] 1.45-3.15) and the incidence density 956 per 100,000 transplant years of follow-up (95% CI 580-1321). Multivariate risk factor analysis and backwards elimination showed a significant positive association between mycophenolate mofetil (MMF), hyposplenism/asplenia, and IPD, whereas trimethoprim-sulfamethoxazole (TMP/SMX) prophylaxis for Pneumocystis jirovecii pneumonia (PJP) was associated with lower odds of IPD cases. Of alloHSCT recipients with IPD, 38.5% required intensive care, and, of deaths documented in cases over the period of review, 30% were attributable to IPD. Serotypes causing IPD matched currently available vaccines in 15/22 (68.1%) episodes. CONCLUSIONS: The incidence of IPD in alloHSCT recipients is an important cause of morbidity and mortality, with rates of disease being many fold higher than the general population. Patients with evidence of hyposplenism/asplenia define a high-risk group in the alloHSCT population for IPD, and the independent association with IPD and MMF in the adjusted model from this study requires further evaluation. The occurrence of post-transplant IPD may be reduced by measures such as vaccination with both 13-valent and 23-valent pneumococcal vaccines. TMP/SMX prophylaxis for the prevention of PJP may offer incidental protection against IPD in alloHSCT recipients.

Site directed immobilization of glucose-6-phosphate dehydrogenase via thiol-disulfide interchange: influence on catalytic activity of cysteines introduced at different positions.

This study shows the effect of site-directed enzyme immobilization upon the enzyme activity of covalently bound glucose-6-phosphate dehydrogenase from Leuconostoc mesenteroides. Immobilization points were introduced at sterically accessible sites in order to control the protein's orientation and twice as much activity was recovered in comparison to conventionally immobilized enzyme. Immobilization of G6PDH via genetically engineered cysteine provided a simple, but effective method to control the immobilization process. G6PDH variants with cysteine close to the active center (L218C), close to the dimer interface (D205C) as well as far from the active center (D453C) showed changes in activity and the efficacy of immobilization.

Guidelines for the prevention of sepsis in asplenic and hyposplenic patients.

Asplenic or hyposplenic patients are at risk of fulminant sepsis. This entity has a mortality of up to 50%. The spectrum of causative organisms is evolving as are recommended preventive strategies, which include education, prophylactic and standby antibiotics, preventive immunizations, optimal antimalarial advice when visiting endemic countries and early management of animal bites. However, there is evidence that adherence to these strategies is poor. Consensus-updated guidelines have been developed to help Australian and New Zealand clinicians and patients in the prevention of sepsis in asplenic and hyposplenic patients.

How far does a doctor's 'duty of care' go?

It is a long-standing tradition in medicine that doctors have an ethical duty to care for all patients who fall within the scope of their skill base. This duty reflects the value system of many doctors and the type of typical dedication to their craft that has long been expected and given. The modern doctor, however, may have other additional roles -- such as those of parent, researcher, business person and many others. What about the duties that accompany these other activities and what if these duties come into conflict with the duty to care for patients? How does a doctor decide how far the duty to care for patients extends? This article explores this question of duty and discusses how the notion of the traditional doctor's duty to care may need to be amended in light of the kinds of lives that doctors now lead.

Toxoplasmosis. Are cats really the source?

BACKGROUND: Toxoplasmosis is a zoonotic infection caused by the parasite Toxoplasma gondii. It would appear to be one of the commonest human infections, but is usually asymptomatic. It is only problematic if the patient is pregnant or immunosuppressed. OBJECTIVE: To discuss the epidemiology, clinical features, diagnosis, treatment and prevention of toxoplasmosis. DISCUSSION: Cats have long been blamed for playing a major role as the reservoir of this infection. To some extent, this is true, although contact with infected raw meat is probably a more important cause of human infection in many countries. Direct contact with pet cats is also a less common risk factor for infection than ingestion of the oocyst from faecally contaminated hands (often via soil).

Amino acid similarity matrices based on force fields.

MOTIVATION: We propose a general method for deriving amino acid substitution matrices from low resolution force fields. Unlike current popular methods, the approach does not rely on evolutionary arguments or alignment of sequences or structures. Instead, residues are computationally mutated and their contribution to the total energy/score is collected. The average of these values over each position within a set of proteins results in a substitution matrix. RESULTS: Example substitution matrices have been calculated from force fields based on different philosophies and their performance compared with conventional substitution matrices. Although this can produce useful substitution matrices, the methodology highlights the virtues, deficiencies and biases of the source force fields. It also allows a rather direct comparison of sequence alignment methods with the score functions underlying protein sequence to structure threading. AVAILABILITY: Example substitution matrices are available from http://www.rsc.anu.edu.au/~zsuzsa/suppl/matrices.html. SUPPLEMENTARY INFORMATION: The list of proteins used for data collection and the optimized parameters for the alignment are given as supplementary material at http://www.rsc.anu.edu.au/~zsuzsa/suppl/matrices.html.

The pathology of human and murine pulmonary infection with Cryptococcus neoformans var. gattii.

Human infection by Cryptococcus neoformans var. neoformans is well characterised and usually occurs in immunocompromised patients. Less is known about infection by Cryptococcus neoformans var. gattii, which usually produces disease in previously normal individuals. In two cases of human pulmonary infection by Cryptococcus neoformans var. gattii, we observed a mixed inflammatory pattern, including granulomas associated with numerous T lymphocytes and a lymphocytic interstitial pneumonitis with B lymphocytes and formation of follicles. We also established a murine model of pulmonary infection by Cryptococcus neoformans var. gattii, which reproduced most of these features. This model is likely to prove useful in studies of the pathogenesis of this infection.

Automated protein design and sequence optimisation: scoring functions and the search problem.

Advances in molecular biology may mean that almost any protein sequence can be synthesised, but perhaps this has served to highlight the inadequacy of theoretical work. For a given protein fold, it is probably not possible to reliably predict an "ideal" sequence. We identify and survey several aspects of the problem. Firstly, it is not clear what is the best way to score a sequence-structure pair. Secondly, there is no consensus as to what the score function should represent (free energy or some abstract measure of sequence-structure compatibility). Finally, the number of possible sequences is astronomical and searching this space poses a daunting optimisation problem. These problems are discussed in the light of recent experimental successes.

Protein fold recognition score functions: unusual construction strategies.

We describe two ways of optimizing score functions for protein sequence to structure threading. The first method adjusts parameters to improve sequence to structure alignment. The second adjusts parameters so as to improve a score function's ability to rank alignments calculated in the first score function. Unlike those functions known as knowledge-based force fields, the resulting parameter sets do not rely on Boltzmann statistics, have no claim to representing free energies and are purely constructions for recognizing protein folds. The methods give a small improvement, but suggest that functions can be profitably optimized for very specific aspects of protein fold recognition. Proteins 1999;36:454-461.

Enhanced protein fold recognition using secondary structure information from NMR.

NMR offers the possibility of accurate secondary structure for proteins that would be too large for structure determination. In the absence of an X-ray crystal structure, this information should be useful as an adjunct to protein fold recognition methods based on low resolution force fields. The value of this information has been tested by adding varying amounts of artificial secondary structure data and threading a sequence through a library of candidate folds. Using a literature test set, the threading method alone has only a one-third chance of producing a correct answer among the top ten guesses. With realistic secondary structure information, one can expect a 60-80% chance of finding a homologous structure. The method has then been applied to examples with published estimates of secondary structure. This implementation is completely independent of sequence homology, and sequences are optimally aligned to candidate structures with gaps and insertions allowed. Unlike work using predicted secondary structure, we test the effect of differing amounts of relatively reliable data.

Sausage: protein threading with flexible force fields.

Sausage is a protein sequence threading program, but with remarkable run-time flexibility. Using different scripts, it can calculate protein sequence-structure alignments, search structure libraries, swap force fields, create models from alignments, convert file formats and analyse results. There are several different force fields which might be classed as knowledge-based, although they do not rely on Boltzmann statistics. Different force fields are used for alignment calculations and subsequent ranking of calculated models.

The dependence of amino acid pair correlations on structural environment.

A statistical analysis was performed to determine to what extent an amino acid determines the identity of its neighbors and to what extent this is determined by the structural environment. Log-linear analysis was used to discriminate chance occurrence from statistically meaningful correlations. The classification of structures was arbitrary, but was also tested for significance. A list of statistically significant interaction types was selected and then ranked according to apparent importance for applications such as protein design. This showed that, in general, nonlocal, through-space interactions were more important than those between residues near in the protein sequence. The highest ranked nonlocal interactions involved residues in beta-sheet structures. Of the local interactions, those between residues i and i + 2 were the most important in both alpha-helices and beta-strands. Some surprisingly strong correlations were discovered within beta-sheets between residues and sites sequentially near to their bridging partners. The results have a clear bearing on protein engineering studies, but also have implications for the construction of knowledge-based force fields.

Protein fold recognition without Boltzmann statistics or explicit physical basis.

We present a fast method for finding optimal parameters for a low-resolution (threading) force field intended to distinguish correct from incorrect folds for a given protein sequence. In contrast to other methods, the parameterization uses information from >10(7) misfolded structures as well as a set of native sequence-structure pairs. In addition to testing the resulting force field's performance on the protein sequence threading problem, results are shown that characterize the number of parameters necessary for effective structure recognition.

Necrotizing cutaneous infection caused by Curvularia brachyspora in an immunocompetent host.

A case of necrotizing cutaneous infection in an immunocompetent host caused by the dematiaceous fungus, Curvularia brachyspora is presented. A 58-year-old man was transferred to the Prince Henry Hospital for treatment of necrotizing cellulitis involving both thighs that developed following cryotherapy. The patient did not respond to antibacterial therapy and hyperbaric oxygen. He was empirically commenced on intravenous amphotericin B and began to respond even before the pathogen C. brachyspora was identified.

Perspectives in protein-fold recognition.

Fold recognition force fields based on statistics from native structures have become commonplace. New, nonphysical force fields based on optimizing parameters rather than reflecting Boltzmann statistics may offer improvement in force-field performance for threading and other applications. Improvements in sequence-to-structure alignments will also be essential for improved fold recognition.

Dynamical studies of peptide motifs in the Plasmodium falciparum circumsporozoite surface protein by restrained and unrestrained MD simulations.

The immunodominant region on the circumsporozoite surface (CS) protein of the malaria parasite Plasmodium falciparum contains 37 repeated copies of a asparagine-alanine-asparagine-proline (NANP) motif NMR studies of linear synthetic peptides containing one, two or three repeat units provided evidence for nascent type I beta-turns within the NPNA cadence in aqueous solution. The beta-turns could be stabilised upon substituting proline for alpha-methylproline (p(Me)) in the dodecamer (NP(Me)NA)3, without loss of the ability to elicit antibodies cross-reactive with P. falciparum sporozoites. In this work, four 4 ns MD simulations of the dodecapeptide Acetyl-(NP(Me)NA)3, in water, using NOE distance restraints, using 3J-coupling constant restraints, using both these restraints and without restraints, were carried out to determine the conformations of this peptide in aqueous solution. An unrestrained MD simulation of the unmethylated Ac-(NPNA)3 peptide in water was also carried out to investigate the effect of the additional methyl groups on the structure and dynamics of the peptide. The application of NOE distance restraints and 3J-coupling constant restraints leads to contradictory results, probably due to different averaging time scales inherent to the measurement of these data, which exceed the 100 ps averaging applied in the simulations. The additional methyl groups lead to more compact structures, which display enhanced local fluctuations. The central tetrapeptide adopts a type I beta-turn, while the outer motifs display more conformational variability. The three motifs in the methylated dodecamer peptide, however, adopt frequently in the distance restrained MD simulation a compact structure such that the outer motifs appear to form a hydrophobic core by stacking of their two proline rings. This arrangement also suggests how a peptide containing multiple tandemly linked copies of a stable beta-turn NPNA motif might adopt a folded stem-like structure, which conceivably may be of biological relevance in the native CS protein.

Protein structure prediction force fields: parametrization with quasi-newtonian dynamics.

We present an unusual method for parametrizing low-resolution force fields of the type used for protein structure prediction. Force field parameters were determined by assigning each a fictitious mass and using a quasi-molecular dynamics algorithm in parameter space. The quasi-energy term favored folded native structures and specifically penalized folded nonnative structures. The force field was generated after optimizing less than 70 adjustable parameters, but shows a strong ability to discriminate between native structures and compact misfolded alternatives. The functional form of the force field was chosen as in molecular mechanics and is not table-driven. It is continuous with continuous derivatives and is thus suitable for use with algorithms such as energy minimization or newtonian dynamics.

Molecular dynamics simulation using weak-coupling NOE distance restraining.

Application of the weak-coupling scheme to restrain the configurations of a molecular system to a set of NOE distance restraints is investigated using two test systems: (i) a 15-atom chain molecule with one distance restraint; and (ii) a protein molecule with hundreds of NOE distance restraints. Atom-atom distance restraining by the weak-coupling technique is possible, but this method does not produce as good results as the penalty function method normally used to maintain NOE distance restraints.