Microbial resistance: what is to be done? Information from the panel of experts

This document represents the recommendations of a panel of Spanish experts on antibiotic use and resistance. In a Task Force, under the auspices of the Spanish Ministry of Health and Consumer Affairs that took place in 1994 in Madrid, the members were gravely concerned about the national increase in antibiotic resistance. They analysed the development, evolution and spread of antibiotic resistance among community-acquired human bacterial pathogens in Spain, its relation with antibiotic consumption, and they proposed future surveillance strategies for monitoring the patterns of antibiotic use and consumption. Success will require a collective action among the producers (pharmaceutical industry), prescribers (doctors, veterinarians), dispensers (pharmacists), and consumers (patients). Two similar documents have been recently published by the American Society of Microbiology and the World Health Organization showing the global concern about this topic.     

The effect of superoxide dismutase on the recovery of human bone marrow hemopoietic stem cells stored at 4 degreesC

Epidemiological research during the past 40 years has demonstrated with increasing clarity that amphibole asbestos fibres--crocidolite, amosite and tremolite--are more carcinogenic than chrysotile. A smaller number of well-controlled studies using lung burden analyses, while adding to the specificity of this conclusion, have shown that amphibole fibres also differ from chrysotile in being far more durable and biopersistent in lung tissue. Analyses of mesothelioma and lung cancer in a large cohort of Canadian chrysotile miners and millers have recently shown that the low-level presence of fibrous tremolite in these mines, rather than the chrysotile, may well be responsible. The high risk of lung cancer, but not of mesothelioma, in the chrysotile textile industry remains anomalous and cannot be explained in this way. These various findings are directly relevant to the choice of the experimental methods which should be used for screening man-made fibres for industrial use. Although it is clear that biopersistence is a major determinant of cancer risk in animals, and perhaps also in man, other factors affecting the biological activity of mineral fibres may also be important.     

Brachial plexus injury during cardiac catheterisation in children. Report of two cases

The arm is particularly vulnerable to neurological injury, due to the intimate anatomical relationship between the brachial plexus and the traction zones. Severe injuries of the brachial plexus can be caused by compression, traction or laceration. Fortunately, many deficits are superficial and permanent neurovascular deficits are rare. Nevertheless, it is important to identify the probable cause of the injury since the prognosis for recovery directly depends on the underlying nature of the neurological deficit. Two anaesthetised children who suffered brachial plexus injury during cardiac catheterisation are reported. The first, with Ebstein's anomaly and significant pulmonary valve stenosis, presented, after the procedure, with a right arm motor deficit with proximal predominance. The second patient, with tetralogy of Fallot and pulmonary atresia, presented difficulty in left arm abduction and external rotation on awakening. The risk factors for brachial plexus lesions during anaesthesia are discussed. These include improper positioning, anaesthetic agents, extreme variations of body mass index and anatomical anomalies. Prevention, evolution and treatment of the brachial plexus injury are also considered. With proper care by the cardio-radiologist and anaesthesiologist the frequency of this injury can be reduced.     

In vivo studies on the role of the peripheral benzodiazepine receptor (PBR) in steroidogenesis

In various steroidogenic cell models, mitochondrial preparations and submitochondrial fractions, the expression of the mitochondrial 18 kDa peripheral-type benzodiazepine receptor (PBR) protein confers the ability to take up and release, upon ligand activation, cholesterol. Thus, cholesterol becomes available to P450scc on the inner mitochondrial membrane. These in vitro studies were validated by in vivo experiments. Treatment of rats with ginkgolide B (GKB), specifically reduced the ligand binding capacity, protein, and mRNA expression of the adrenocortical PBR and circulating glucocorticoid levels. Treatment with GKB also resulted in inhibition of PBR protein synthesis and corticosterone production by isolated adrenocortical cells in response to ACTH. The ontogeny of both PBR binding capacity and protein directly paralleled that of ACTH-inducible steroidogenesis in rat adrenal cells and in rats injected with ACTH. In addition, the previously described suppression of luteal progesterone synthesis in the pregnant rat by continuous in vivo administration of a gonadotropin-releasing hormone agonist may be due to decreased luteal PBR ligand binding and mRNA. These results suggest that (i) PBR is an absolute prerequisite for adrenocortical and luteal steroidogenesis, (ii) regulation of adrenal PBR expression may be used as a tool to control circulating glucocorticoid levels and (iii) the stress hypo-responsive period of neonatal rats may result from decreased adrenal cortical PBR expression.     

The FML (Fucose Mannose Ligand) of Leishmania donovani. a new tool in diagnosis, prognosis, transfusional control and vaccination against human kala-azar

Upon fractionation of a post mitochondrial supernatant from rat liver, phosphorylase kinase activity was largely recovered in the cytosol and the smooth endoplasmic reticulum (SER) fraction. The presence of phosphorylase kinase in SER vesicles was not due to an interaction of the enzyme with glycogen particles, since previous elimination of SER glycogen either by 48 h animal starvation or by treatment of the membrane fraction with alpha-amylase did not significantly alter phosphorylase kinase activity content. Washing of the initial pellet of SER fraction (crude SER) by dilution and recentrifugation, released in the supernatant an amount of phosphorylase kinase activity, which is dependent on: i) the degree of dilution, ii) the number of washes, iii) the ionic strength of the washing solution and iii) the presence or absence of Ca2+. Crude SER-associated phosphorylase kinase was marginally affected by increased concentrations of antibody against rabbit skeletal muscle holoenzyme which nevertheless drastically inhibited cytosolic enzyme activity, while it showed a higher resistance to partial proteolysis and a different Western blotting profile with anti-phosphorylase kinase when compared with the soluble kinase. A small but significant fraction of SER phosphorylase kinase was strongly associated with the microsomal fraction being partly extractable only in presence of detergents. This membrane-bound enzyme form exhibited an alkaline pH optimum, in contrast to the neutral pH optima of both soluble and weakly associated phosphorylase kinase.     

Characteristics of multicellular spheroids formed by primary cultures of human thyroid tumor cells

Features of multicellular tumor spheroids (MCTS) differed depending on their types of cells. MCTS formed by 4000 human thyroid primary culture epithelial tumor cells displayed diameters between 0.31 and 0.33 mm within 2 days regardless of the stage of malignancy of the originating tumors. Their cellular composition reflected that of the originating tumor in regard to DNA content and the expression of cytokeratin, vimentin, as well as thyroglobulin. During the following 3 weeks, their sizes increased up to diameters of 0.42 mm when their cells had been derived from carcinomas, and MCTS originating from adenomas stopped growing within the next 2 days. After 8 days of incubation, proliferating cells were only found in carcinoma MCTS. The cells were randomly distributed over the total volume of the spheroids, which displayed irregular cell arrangements but not concentric cell layers and did not form necrotic centers.     

Mutational analysis of the Vibrio fischeri LuxI polypeptide: critical regions of an autoinducer synthase

Synthesis of the Vibrio fischeri autoinducer, a signal involved in the cell density-dependent activation of bioluminescence, is directed by the luxI gene product. The LuxI protein catalyzes the synthesis of N-acyl-homoserine lactones from S-adenosylmethionine and acylated-acyl carrier protein. We have gained an appreciation of the LuxI regions and amino acid residues involved in autoinducer synthesis by isolating and analyzing mutations generated by random and site-specific mutagenesis of luxI. By random mutagenesis we isolated 13 different single amino acid substitutions in the LuxI polypeptide. Eleven of these substitutions resulted in no detectable autoinducer synthase activity, while the remaining two amino acid substitutions resulted in reduced but detectable activity. The substitutions that resulted in no detectable autoinducer synthase activity mapped to two small regions of LuxI. In Escherichia coli, wild-type luxI showed dominance over all of the mutations. Because autoinducer synthesis has been proposed to involve formation of a covalent bond between an acyl group and an active-site cysteine, we constructed site-directed mutations that altered each of the three cysteine residues in LuxI. All of the cysteine mutants retained substantial activity as an autoinducer synthase in E. coli. Based on the analysis of random mutations we propose a model in which there are two critical regions of LuxI, at least one of which is an intimate part of an active site, and based on the analysis of site-directed mutations we conclude that an active-site cysteine is not essential for autoinducer synthase activity.