Detection of meticillin-resistant Staphylococcus aureus and carbapenemase-producing Enterobacteriaceae in Danish emergency departments - evaluation of national screening guidelines.

BACKGROUND: Multi-resistant bacteria (MRB) are an emerging problem. Early identification of patients colonized with MRB is mandatory to avoid in-hospital transmission and to target antibiotic treatment. Since most patients pass through specialized emergency departments (EDs), these departments are crucial in early identification. The Danish National Board of Health (DNBH) has developed exposure-based targeted screening tools to identify and isolate carriers of meticillin-resistant Staphylococcus aureus (MRSA) and carbapenemase-producing Enterobacteriaceae (CPE). AIM: To assess the national screening tools for detection of MRSA and CPE carriage in a cohort of acute patients. The objectives were to investigate: (i) if the colonized patients were detected; and (ii) if the colonized patients were isolated. METHODS: This was a multi-centre cross-sectional survey of adults visiting EDs. The patients answered the DNBH questions, and swabs were taken from the nose, throat and rectum. The collected samples were examined for MRSA and CPE. Screening performances were calculated. FINDINGS: Of the 5117 included patients, 16 were colonized with MRSA and four were colonized with CPE. The MRSA screening tool had sensitivity of 50% [95% confidence interval (CI) 25-75%] for carrier detection and 25% (95% CI 7-52%) for carrier isolation. The CPE screening tool had sensitivity of 25% (95% CI 1-81%) and none of the CPE carriers were isolated. CONCLUSION: The national screening tools were of limited use as the majority of MRSA and CPE carriers passed unidentified through the EDs, and many patients were isolated unnecessarily.

Local infiltration analgesia for total knee arthroplasty: should ketorolac be added?

BACKGROUND: Adequate postoperative analgesia with minimal side-effects is essential for early mobilization and recovery in patients undergoing total knee arthroplasty (TKA). High-volume local infiltration analgesia (LIA) with ropivacaine has been introduced, but effects of adjuvants are still debated. We tested the hypothesis that the addition of ketorolac to LIA significantly improves analgesia after TKA. METHODS: Sixty patients undergoing TKA were randomized to receive intraoperative LIA (ropivacaine 300 mg and epinephrine 0.5 mg) combined with either ketorolac 30 mg (ketorolac group) or saline (control group). After surgery, eight bolus doses of ropivacaine 100 mg combined with either ketorolac 15 mg (ketorolac group) or saline (control group) were administered every 6 h via an intra-articular catheter. The primary outcome was postoperative consumption of i.v. morphine patient-controlled analgesia (PCA). Secondary outcomes were time to first request of i.v. morphine PCA, pain intensity, side-effects, and readiness for hospital discharge. RESULTS: Consumption of i.v. morphine PCA was lower in the ketorolac group vs control group {0-6 h: 0 (0-0) vs 5 (0-10) mg, P<0.0001; 0-48 h: 10 (0-22.5) vs 48.75 (30-82.5) mg, P<0.0001 [median (inter-quartile range, IQR)]}. Time to first request of i.v. morphine PCA was longer in the ketorolac group vs the control group [490 (248-617) vs 223 (115-319) min, P=0.02, median (IQR)]. Early postoperative pain (<48 h) and readiness for hospital discharge were also significantly reduced in the ketorolac group. CONCLUSIONS: LIA with ketorolac results in reduced morphine consumption, reduced pain intensity, and earlier readiness for hospital discharge.

Consensus on the use of neurophysiological tests in the intensive care unit (ICU): electroencephalogram (EEG), evoked potentials (EP), and electroneuromyography (ENMG).

STUDY AIM: To provide a consensus of European leading authorities about the optimal use of clinical neurophysiological (CN) tests (electroencephalogram [EEG]; evoked potentials [EP]; electroneuromyography [ENMG]) in the intensive care unit (ICU) and, particularly, about the way to make these tests clinically useful for the management of individual patients. METHODS: This study gathered together several European clinical neurophysiologists and neurointensivists whose leading contributions in the adult or paediatric ICU and in continuous neuromonitoring had been peer-acknowledged. It was based on both a literature review and each participant's own experience. Given the methodological impossibility to gather studies fulfilling criteria of evidence-based medicine, this article essentially relies on expert opinions that were gained after several rounds, in which each expert was invited to communicate his own contribution to all other experts. A complete consensus has been reached when submitting the manuscript. RESULTS: What the group considered as the best classification systems for EEG and EP abnormalities in the ICU is first presented. CN tests are useful for diagnosis (epilepsy, brain death, and neuromuscular disorders), prognosis (anoxic ischemic encephalopathy, head trauma, and neurologic disturbances of metabolic and toxic origin), and follow-up, in the adult, paediatric, and neonatal ICU. Regarding prognosis, a clear distinction is made between these tests whose abnormalities are indicative of an ominous prognosis and those whose relative normalcy is indicative of a good prognosis. The prognostic significance of any test may vary as a function of coma etiology. CONCLUSION: CN provides quantitative functional assessment of the nervous system. It can be used in sedated or curarized patients. Therefore, it should play a major role in the individual assessment of ICU patients.

Clinical evaluation of excitability measures in sensory nerve.

A recently described method for recording multiple excitability parameters of human motor nerves has been adapted to the study of sensory nerves. The protocol measures stimulus-response behavior using two stimulus durations (from which the distribution of strength-duration time constants is estimated), threshold electrotonus to 100 ms polarizing currents, a current-threshold relationship (indicating inward and outward rectification), and the recovery of excitability following supramaximal activation. The method was tested on 50 healthy volunteers, stimulating the median nerve at the wrist and recording the antidromic compound sensory nerve action potential (SNAP) from digit 2. The excitability measurements were similar, where comparisons were possible, with published sensory nerve data, and confirmed differences from motor nerves, particularly in strength-duration behavior and recovery cycle, likely to reflect functional differences between sensory and motor nerves. Although slower than for motor nerves, the sensory nerve recordings were sufficiently quick (16 to 18 min) to allow them to be included in routine clinical studies. We propose that this method, which provides quite different and complementary information about nerve function to conventional conduction studies, provides a useful new approach for exploring the pathophysiology of sensory neuropathies.

[Teaching and counseling families with small children in an urban area about minor illnesses of childhood]. / Undervisning og rådgivning af småbørnsfamilier i et storbyområde om småbørns almindelige sygelighed.

INTRODUCTION: Parents' knowledge of and experience with illness in their children are often limited. They find it difficult to take the responsibility when their children are ill. They seek help immediately from the health care system, although the symptoms are mild. We wanted to discover whether teaching parents about minor illness in children and counselling over the telephone during acute illness would mean that parents sought help less often from the health care system, when their children became ill. This article deals with the teaching of the parents. MATERIAL AND METHODS: The project had an intervention group of 91 children aged 12-35 months and a control group of 400 children aged 9-33 months. The intervention group was instructed for two hours twice in the local day nursery and telephone counselling from a nurse trained in children's acute illnesses was made available from 4 pm to 11 pm on weekdays and from 10 am to 11 pm at weekends during the period 20th October 1998 to 1st May 1999. RESULTS: No visible change was seen in the use of the health care system in either group, but there was a small change in favour of the intervention group in the use of a general practitioner instead of calling the emergency medical service. Eighty per cent of the counselling dealt with common acute illnesses; 75% ended without referral. DISCUSSION: There was no visible difference in the use of the health care system in either group. Parents were happy with the offer of teaching and telephone counselling, but it should have been given when the infants started at the nursery. They felt their children were too old at the time the project started. We suggest a continuation of antenatal classes in the form of "parent preparation classes", where one subject could be minor acute illnesses in children.

[Why do parents of small children visit casualty ward when their children suffer of acute illness?]. / Hvorfor opsøger børnefamilier skadestuen ved akut sygdom?

INTRODUCTION: People in big cities tend to use the casualty ward instead of their own doctor or the emergency service. This is both more expensive and an inappropriate use of the health care system in Denmark. We wanted to find out why parents used the casualty ward and to look for an alternative way of helping parents during minor acute illnesses in their children. MATERIAL AND METHOD: One hundred questionnaires were distributed in the paediatric ward of Rigshospitalet, Copenhagen, from 28.1.1999 to 9.4.1999 to parents who came to the casualty ward without a referral from a doctor. Only people who could speak Danish participated. RESULTS: Seventy-five percent had no referral from a doctor, the rest were verbally referred by a doctor. Twenty-five percent came to the casualty ward while their own doctor was on duty. Thirty-nine percent chose the casualty ward, because the staff had a special knowledge of children. Most parents wanted to have their children examined by a paediatrician. DISCUSSION: Parents need to be educated in the proper use of the health care system when their children are ill. Many use the casualty ward for convenience and to get expert knowledge about their children's illness, even when it is not necessary. We suggest that an experienced children's nurse should work with the emergency service, so as to keep children with minor, acute illness out of the casualty ward.

[Ischemic cerebral apoplexy in children]. / Iskaemisk apoplexia cerebri hos børn.

During two years we identified eight children aged 1.5-10 years with cerebral ischaemic stroke. Prior to the stroke seven of eight children were in full health. Predisposing factors were endocarditis and trauma. Four children had prodomal symptoms prior to the infarction. All had acute hemiparesis on admission. Three of the children had fever, and five had varicellazoster infection three to 18 months prior to the stroke. Three children had convulsions. Seven of eight children had stenoses or occlusions of the middle cerebral or the basilar artery. Five children have persistent deficits and none have died. The children did not have coagulopathies, hypertension or arteriosclerosis. Echocardiography did not show patent foramen ovale. Early clinical and neuroradiological investigations are of importance in reaching the appropriate diagnosis.

Multiple measures of axonal excitability: a new approach in clinical testing.

From measurements of nerve excitability and the changes in excitability produced by nerve impulses and conditioning currents, it is possible to infer information about the membrane potential and biophysical properties of peripheral axons. Such information cannot be obtained from conventional nerve conduction studies. This article describes a new method that enables several such measurements to be made on a motor nerve quickly and reproducibly, with minimal operator intervention. The protocol measures stimulus-response behavior using two stimulus durations (from which the distribution of strength-duration time constants can be estimated), threshold electrotonus to 100-ms polarizing currents, a current-threshold relationship (indicating inward and outward rectification), and the recovery of excitability following supramaximal activation. The method was tested on 30 healthy volunteers, stimulating the median nerve at the wrist and recording from the abductor pollicis brevis. The results were comparable with previously published normal data, but the recordings took less than 10 min. The convenience and brevity of the new method make it appropriate for routine clinical use.

Electrostatics in the active site of an alpha-amylase.

The importance of electrostatics in catalysis has been emphasized in the literature for a large number of enzymes. We examined this hypothesis for the Bacillus licheniformis alpha-amylase by constructing site-directed mutants that were predicted to change the pKa values of the catalytic residues and thus change the pH-activity profile of the enzyme. To change the pKa of the catalytic residues in the active site, we constructed mutations that altered the hydrogen bonding network, mutations that changed the solvent accessibility, and mutations that altered the net charge of the molecule. The results show that changing the hydrogen bonding network near an active site residue or changing the solvent accessibility of an active site residue will very likely result in an enzyme with drastically reduced activity. The differences in the pH-activity profiles for these mutants were modest. pH-activity profiles of mutants which change the net charge on the molecule were significantly different from the wild-type pH-activity profile. The differences were, however, difficult to correlate with the electrostatic field changes calculated. In several cases we observed that pH-activity profiles shifted in the opposite direction compared to the shift predicted from electrostatic calculations. This strongly suggests that electrostatic effects cannot be solely responsible for the pH-activity profile of the B. licheniformis alpha-amylase.

Improving macromolecular electrostatics calculations.

Electrostatic interactions play a key role in many aspects of protein engineering. Consequently, much effort has been put into the design of software for calculating electrostatic fields around macromolecules. We show that optimization of hydrogen bonding networks can improve both the results of pK(a) calculations and the results of electrostatic calculations performed by commonly used programs such as DelPhi. Further optimization can often be achieved by flipping the side chains of asparagine, histidine and glutamine around their chi2, chi2 and chi3 torsion angles, respectively, when this improves the local hydrogen bonding network. These optimizations are applied to some well characterized proteins: BPTI, hen egg white lysozyme and superoxide dismutase. A search for flipped residues in the PDB revealed that significant improvements in electrostatic calculations in or near the active site of enzymes can be expected for about one quarter of all enzymes in the PDB.

[X-linked recessive bulbospinal neuronopathy--Kennedy's syndrome]. / X-bundet recessiv bulbospinal neuronopati--Kennedys syndrom.

Kennedy's syndrome is an inherited disease which was probably first described 100 years ago. Although rare, a recent report suggests that the prevalence may show considerable regional differences. A review of 30 different names of the disease is given. Originally, the disorder was regarded as a spinal and bulbar muscular atrophy but it is now obvious that there is severe axonal degeneration, also of the sensory fibres with the pattern of a central-peripheral distal axonal neuropathy. This was also present in the two recognized cases presented here. The sensory symptoms develop slowly and it is suggested that a peripheral sprouting compensates for the loss not only of motor, but also sensory fibres. It is important to distinguish the disease from motor neuron diseases since the progression is slow and the expected life span is normal. The clinical presentation with facial palsy and perioral contraction-fasciculations is pathognomonic. However, demonstration of increased (CAG)n repeat size in the androgen receptor gene is diagnostic. A normal (CAG)n repeat size excludes the diagnosis, since the abnormal expansion is the only mutation associated with the disease. Other types of mutations in the androgen receptor gene lead to a different clinical presentation, testicular feminization.

Analysis of the dynamics of rhizomucor miehei lipase at different temperatures.

The dynamics of Rhizomucor miehei lipase has been studied by molecular dynamics simulations at temperatures ranging from 200-500K. Simulations carried out in periodic boundary conditions and using explicit water molecules were performed for 400 ps at each temperature. Our results indicate that conformational changes and internal motions in the protein are significantly influenced by the temperature increase. With increasing temperature, the number of internal hydrogen bonds decreases, while surface accessibility, radius of gyration and the number of residues in random coil conformation increase. In the temperature range studied, the motions can be described in a low dimensional subspace, whose dimensionality decreases with increasing temperature. Approximately 80% of the total motion is described by the first (i) 80 eigenvectors at T=200K, (ii) 30 eigenvectors at T=300K and (iii) 10 eigenvectors at T=400K. At high temperature, the alpha-helix covering the active site in the native Rhizomucor miehei lipase, the helix at which end the active site is located, and in particular, the loop (Gly35-Lys50) show extensive flexibility.

Conserved residues and their role in the structure, function, and stability of acyl-coenzyme A binding protein.

In the family of acyl-coenzyme A binding proteins, a subset of 26 sequence sites are identical in all eukaryotes and conserved throughout evolution of the eukaryotic kingdoms. In the context of the bovine protein, the importance of these 26 sequence positions for structure, function, stability, and folding has been analyzed using single-site mutations. A total of 28 mutant proteins were analyzed which covered 17 conserved sequence positions and three nonconserved positions. As a first step, the influence of the mutations on the protein folding reaction has been probed, revealing a folding nucleus of eight hydrophobic residues formed between the N- and C-terminal helices [Kragelund, B. B., et al. (1999) Nat. Struct. Biol. (In press)]. To fully analyze the role of the conserved residues, the function and the stability have been measured for the same set of mutant proteins. Effects on function were measured by the extent of binding of the ligand dodecanoyl-CoA using isothermal titration calorimetry, and effects on protein stability were measured with chemical denaturation followed by intrinsic tryptophan and tyrosine fluorescence. The sequence sites that have been conserved for direct functional purposes have been identified. These are Phe5, Tyr28, Tyr31, Lys32, Lys54, and Tyr73. Binding site residues are mainly polar or charged residues, and together, four of these contribute approximately 8 kcal mol-1 of the total free energy of binding of 11 kcal mol-1. The sequence sites conserved for stability of the structure have likewise been identified and are Phe5, Ala9, Val12, Leu15, Leu25, Tyr28, Lys32, Gln33, Tyr73, Val77, and Leu80. Essentially, all of the conserved residues that maintain the stability are hydrophobic residues at the interface of the helices. Only one conserved polar residue, Gln33, is involved in stability. The results indicate that conservation of residues in homologous proteins may result from a summed optimization of an effective folding reaction, a stable native protein, and a fully active binding site. This is important in protein design strategies, where optimization of one of these parameters, typically function or stability, may influence any of the others markedly.

Structure of the Bacillus agaradherans family 5 endoglucanase at 1.6 A and its cellobiose complex at 2.0 A resolution.

The enzymatic degradation of cellulose, by cellulases, is not only industrially important in the food, paper, and textile industries but also a potentially useful method for the environmentally friendly recycling of municipal waste. An understanding of the structural and mechanistic requirements for the hydrolysis of the beta-1,4 glycosidic bonds of cellulose is an essential prerequisite for beneficial engineering of cellulases for these processes. Cellulases have been classified into 13 of the 62 glycoside hydrolase families [Henrissat, B., and Bairoch, A. (1996) Biochem J. 316, 695-696]. The structure of the catalytic core of the family 5 endoglucanase, Ce15A, from the alkalophilic Bacillus agaradherans has been solved by multiple isomorphous replacement at 1.6 A resolution. Ce15A has the (alpha/beta)8 barrel structure and signature structural features typical of the grouping of glycoside hydrolase families known as clan GH-A, with the catalytic acid/base Glu 139 and nucleophile Glu 228 on barrel strands beta 4 and beta 7 as expected. In addition to the native enzyme, the 2.0 A resolution structure of the cellobiose-bound form of the enzyme has also been determined. Cellobiose binds preferentially in the -2 and -3 subsites of the enzyme. Kinetic studies on the isolated catalytic core domain of Ce15A, using a series of reduced cellodextrins as substrates, suggest approximately five to six binding sites, consistent with the shape and size of the cleft observed by crystallography.

Scoring functions for computational algorithms applicable to the design of spiked oligonucleotides.

Protein engineering by inserting stretches of random DNA sequences into target genes in combination with adequate screening or selection methods is a versatile technique to elucidate and improve protein functions. Established compounds for generating semi-random DNA sequences are spiked oligonucleotides which are synthesised by interspersing wild type (wt) nucleotides of the target sequence with certain amounts of other nucleotides. Directed spiking strategies reduce the complexity of a library to a manageable format compared with completely random libraries. Computational algorithms render feasible the calculation of appropriate nucleotide mixtures to encode specified amino acid subpopulations. The crucial element in the ranking of spiked codons generated during an iterative algorithm is the scoring function. In this report three scoring functions are analysed: the sum-of-square-differences function s, a modified cubic function c, and a scoring function m derived from maximum likelihood considerations. The impact of these scoring functions on calculated amino acid distributions is demonstrated by an example of mutagenising a domain surrounding the active site serine of subtilisin-like proteases. At default weight settings of one for each amino acid, the new scoring function m is superior to functions s and c in finding matches to a given amino acid population.

Impotence and nerve entrapment in long distance amateur cyclists.

OBJECTIVES: To assess the frequency and duration of symptoms suggesting peripheral nerve compression after long distance cycling. MATERIAL AND METHODS: A questionnaire based cross sectional study among 260 participants in a Norwegian annual bicycle touring race of 540 km. RESULTS: Thirty-five of 160 responding males (22%) reported symptoms from the innervation area of the pudendal or cavernous nerves. Thirty-three had penile numbness or hypaesthesia after the tour. In 10, the numbness lasted for more than one week. Impotence was reported by 21 (13%) of the males. It lasted for more than one week in 11, and for more than one month in three. Both genital numbness and impotence were correlated with weakness in the hands after the ride, a complaint reported by 32 (19%) of all 169 respondents. Forty-six cyclists (30%) indicated paraesthesia or numbness in the fingers, half of them from the ulnar nerve area only. CONCLUSION: The frequency of impotence, numbness of the penis, hand weakness and sensory symptoms from the fingers in bicycle sport may be higher than hitherto recognized. It afflicts both experienced cyclists and novices. In some, the complaints may last up to eight months. Besides changing the hand and body position on the bike, restricting the training intensity, and taking ample pauses may also be necessary in prolonged and vigorous bicycle riding to prevent damage to peripheral nerves.