Studies on the cellular defense reactions of the Madeira cockroach, Leucophaea maderae: nodule formation in response to injected bacteria

Nodules were formed in the Madeira cockroach, Leucophaea maderae, in response to injections of low doses (3 x 10(4) bacteria/insect) of three strains of Bacillus cereus and Escherichia coli K12 D31. The most pathogenic strain of bacteria used, B. cereus B1, produced the greatest cellular response, while the least pathogenic, E. coli K12 D31, injected at the same dose, caused little nodule formation. Similarly, nodules were generally found to be larger following injection of pathogenic bacteria such as B. cereus B1 than to the weak pathogen, E. coli K12 D31. There was, however, no difference in the extent of nodule formation with the four bacterial strains/species if they were heat killed prior to injection. Histologically, the nodules formed in response to all bacterial species employed were similar, with a central necrotic core enclosing cell debris and occasional bacteria, and an outer, thin sheath of plasmatocyte-like hemocytes. Possible reasons for the enhanced cellular reactivity observed in L. maderae to pathogenic bacteria are discussed.     

Nucleotide sequence of the Vibrio alginolyticus calcium-dependent, detergent-resistant alkaline serine exoprotease A

The nucleotide sequence of the Vibrio alginolyticus alkaline serine exoprotease A (ProA) gene cloned in Escherichia coli was determined. The exoprotease A gene (proA) consisted of 1602 bp which encoded a protein of 534 amino acids (aa) with an Mr of 55,900. The region upstream from the gene was characterized by a putative promoter consensus region (-10 -35), a ribosome-binding site and ATG start codon. The proA gene encodes a typical 21-aa N-terminal signal sequence which, when fused to alkaline phosphatase by means of transposon TnphoA, was able to mediate transport of the alkaline phosphatase to the periplasm in E. coli. Deletions of up to 106 aa from the C terminus of ProA did not result in the loss of extracellular protease activity. Additional V. alginolyticus genes were not involved in the secretion into the medium of the cloned ProA in E. coli. The amino acid sequence of ProA showed low overall homology to a Serratia marcescens serine exoprotease but significant homology was detected with other subtilisin family exoproteases. The fungal proteinase K, another sodium dodecyl sulfate-resistant protease, had 44% aa homology with ProA.     

Characterization of an extremely thermostable glutamate dehydrogenase: a key enzyme in the primary metabolism of the hyperthermophilic archaebacterium, Pyrococcus furiosus.

Glutamate dehydrogenase (L-glutamate:NAD(P)+ oxidoreductase, deaminating, EC 1.4.1.3) from the hyperthermophilic Archeon Pyrococcus furiosus was purified to homogeneity by chromatography on anion-exchange, molecular-exclusion and hydrophobic-interaction media. The purified native enzyme had an M(r) of 270,000 +/- 15,000 and was shown to be a hexamer with identical subunits of M(r) 46,000. The enzyme was exceptionally thermostable, having a half-life of 3.5 to more than 10 h at 100 degrees C, depending on the concentration of enzyme. The Km of the enzyme for ammonia was high (9.5 mM), indicating that the enzyme is probably active in the deaminating, catabolic direction. The coenzyme utilization of the enzyme resembled the equivalent enzymes from eukaryotes rather than eubacteria, since both NADH and NADPH were recognized with high affinity. The enzyme displayed a preference for NADP+ over NAD+ that was more pronounced at low assay temperatures (50-70 degrees C) compared with the optimal temperature for enzyme activity, 95 degrees C.     

The scl gene product is required for the generation of all hematopoietic lineages in the adult mouse.

Homozygosity for a null mutation in the scl gene causes mid-gestational embryonic lethality in the mouse due to failure of development of primitive hematopoiesis. Whilst this observation established the role of the scl gene product in primitive hematopoiesis, the death of the scl null embryos precluded analysis of the role of scl in later hematopoietic development. To address this question, we created embryonic stem cell lines with a homozygous null mutation of the scl gene (scl-/-) and used these lines to derive chimeric mice. Analysis of the chimeric mice demonstrates that the scl-/- embryonic stem cells make a substantial contribution to all non-hematopoietic tissues but do not contribute to any hematopoietic lineage. These observations reveal a crucial role for the scl gene product in definitive hematopoiesis. In addition, in vitro differentiation assays with scl-/- embryonic stem cells showed that the scl gene product was also required for formation of hematopoietic cells in this system.     

Evolutionary relationships of bacterial and archaeal glutamine synthetase genes

Glutamine synthetase (GS), an essential enzyme in ammonia assimilation and glutamine biosynthesis, has three distinctive types: GSI, GSII and GSIII. Genes for GSI have been found only in bacteria (eubacteria) and archaea (archaebacteria), while GSII genes only occur in eukaryotes and a few soil-dwelling bacteria. GSIII genes have been found in only a few bacterial species. Recently, it has been suggested that several lateral gene transfers of archaeal GSI genes to bacteria may have occurred. In order to study the evolution of GS, we cloned and sequenced GSI genes from two divergent archaeal species: the extreme thermophile Pyrococcus furiosus and the extreme halophile Haloferax volcanii. Our phylogenetic analysis, which included most available GS sequences, revealed two significant prokaryotic GSI subdivisions: GSI-a and GSI-. GSIa-genes are found in the thermophilic bacterium, Thermotoga maritima, the low G+C Gram-positive bacteria, and the Euryarchaeota (includes methanogens, halophiles, and some thermophiles). GSI--type genes occur in all other bacteria. GSI-- and GSI--type genes also differ with respect to a specific 25-amino-acid insertion and adenylylation control of GS enzyme activity, both absent in the former but present in the latter. Cyanobacterial genes lack adenylylation regulation of GS and may have secondarily lost it. The GSI gene of Sulfolobus solfataricus, a member of the Crenarchaeota (extreme thermophiles), is exceptional and could not be definitely placed in either subdivision. The S. solfataricus GSI gene has a shorter GSI--type insertion, but like GSI-a-type genes, lacks conserved sequences about the adenylylation site. We suspect that the similarity of GSI- genes from Euryarchaeota and several bacterial species does not reflect a common phylogeny but rather lateral transmission between archaea and bacteria.Correspondence to: J.R. Brown 1073     

Octopamine receptor subtypes and their modes of action

Octopamine receptor subclasses were first proposed to explain differences in the pharmacological profiles of a range of physiological responses to octopamine obtained in the extensor-tibiae neuromuscular preparation of the locust. Thus, OCTOPAMINE₁ receptors which inhibit an endogenous myogenic rhythm, increase intracellular calcium levels. Also OCTOPAMINE₂ receptors which modulate neuromuscular transmission in this preparation, increase the level of adenylate cyclase activity. The current status of this classification is reviewed by examining the pharmacology of responses to octopamine in a range of preparations. It is concluded that the distinction between OCTOPAMINE₁ and OCTOPAMINE₂ receptor types is still valid, but that OCTOPAMINE₂ receptors exhibit some tissue specific variations. Studies on a clonedDrosophila octopamine/tyramine (phenolamine) receptor are discussed and illustrate many of the difficulties presently encountered in making a definitive classification of octopamine receptors. These include the possibilities that single receptors may activate multiple second messenger systems and that different agonists may differentially couple the same receptor to different second messenger systems.     

University acknowledges special risks,introduces drug program for anesthetists.

Anesthetists are at special risk for becoming addicted to some of the drugs they work with. The problem was highlighted by the recent death of a physician in Comox, BC, and a doctor''s brush with death at a hospital near Ottawa. The University of Ottawa has responded with a program, the professional Assistance Program for the Impaired or Disabled Physician, that is in the final stages of approval. Although it will be aimed initially at anesthetists and anesthesia residents in the university''s teaching hospitals, there are hopes the program will spread eventually to other medical departments.     

Home care for patients with suspected myocardial infarction: use made by general practitioners of a hospital team for initial management.

Two hundred and sixty three general practitioners were offered the use of a hospital based service consisting of a medical senior house officer, a nurse attached to a coronary care unit, and a specially equipped ambulance estate car to help with the initial management of patients with suspected myocardial infarction who might be suitable for home care. One hundred and sixty nine general practitioners registered as potential users of this service; during 22 months they called the hospital team to see 271 patients, 235 of whom the team suspected had indeed suffered a myocardial infarction. During the same period, however, these general practitioners also admitted 317 patients with suspected myocardial infarction directly to hospital. Other general practitioners admitted 323 patients and deputising doctors 258. A further 529 patients with suspected infarction were admitted without the intervention of a general practitioner. Of the patients seen by the team, 54 required immediate admission to hospital; 17 of the remaining patients who initially appeared suitable for home care later required admission to hospital. In a large city such as Nottingham the provision of hospital based facilities to help general practitioners with home management is unlikely to make an appreciable impact on the overall pattern of care of patients with suspected myocardial infarction.