Colonization factors of diarrheagenicE. coli and their intestinal receptors

WhileEscherichia coli is common as a commensal organism in the distal ileum and colon, the presence of colonization factors (CF) on pathogenic strains ofE. coli facilitates attachment of the organism to intestinal receptor molecules in a species- and tissue-specific fashion. After the initial adherence, colonization occurs, and the involvement of additional virulence determinants leads to illness. EnterotoxigenicE. coli (ETEC) is the most extensively studied of the five categories ofE. coli that cause diarrheal disease, and has the greatest impact on health worldwide. ETEC can be isolated from domestic animals and humans. The biochemistry, genetics, epidemiology, antigenic characteristics, and cell and receptor binding properties of ETEC have been extensively described. Another major category, enteropathogenicE. coli (EPEC), has virulence mechanisms, primarily effacement and cytoskeletal rearrangement of intestinal brush borders, that are distinct from ETEC. An EPEC CF receptor has been purified and characterized as a sialidated transmembrane glycoprotein complex directly attached to actin, thereby associating CF-binding with host-cell response. Three, additional categories ofE. coli diarrheal disease, their colonization factors and their host cell receptors are discussed. It appears that biofilms exist in the intestine in a manner similar to oral bacterial biofilms, and thatE. coli is part of these biofilms as both commensals and pathogens.Abbreviations CF colonization factor - CFA Colonization Factor Antigen - CS coli-surface-associated antigen - EAggEC enteroaggregativeE. coli - ECDD E. coli diarrheal disease - EHEC enterohemorrhagicE. coli - EIEC enteroinvasiveE. coli - EPEC enteropathogenicE. coli - ETEC enterotoxigenicE. coli - Gal galactose - GalNAc N-acetyl galactosamine - LT heat-labile toxin - NeuAc N-acetyl neuraminic acid - PCF Putative colonization factor - RBC red blood cells - SLT Shiga-like toxin - ST heat-stable toxin     

Cloning and expression of the Neisseria meningitidis 5C outer membrane protein

The 5C outer membrane protein, one of the N. meningitidis class 5 proteins, was preferably expressed in bacteria isolated from the nasopharynx and its role in adhering to the mucosal cells and invading them as well as the development of anti-5C antibodies in healthy carriers was demonstrated. Anti-5C monoclonal antibodies are bactericidal in the presence of the human complement. The immunodominant region of the 5C protein is highly conserved among the different strains of N. meningitidis, and the opc gene, which encodes the protein, does not seem to show antigenic variations. Here the isolation of the opc gene from the Cuban strain B:4:P1.15 by PCR (Polymerase Chain Reaction) is presented. Under the regulation of the tryptophan promoter, the gene was cloned and sequenced in E. coli with a high level of expression and fused to the amino-terminal end of the interleukin-2 gene. In the dot-blot experiments, the presence of the gene in those strains which did not express the protein in the whole cell ELISA was also detectable.     

The age, growth and reproduction of Chondrostoma polylepis willkommi in a seasonal stream in the Guadalquivir River basin (southern Spain)

The age, growth and reproduction of Chondrostoma polylepis willkommi , an endemic cyprinid from the Iberian Peninsula, was studied for 2 years in a seasonal stream in the Guadalquivir River basin. Annual growth was low. Maximum ages observed were 5+ in males and 7+ in females. There was no significant difference in growth between sexes. Seasonal growth period started in May and continued for 2 to 6 months depending on age. The mean lengths of 1 + and 2+ age-groups decreased once the growth period had finished, probably related to reproductive stress and adverse ecological conditions, and this could explain the occurrence of Rosa Lee's phenomenon. Both sexes matured at the end of their second year of life (1+). There was a significant difference from 1: 1 in the overall sex ratio of 334 males to 464 females. Reproductive period started in March and lasted until May. Ch. p. willkommi was a multiple spawner that released two batches of eggs per female each year. The regression between fecundity ( F ) and fork length (l., mm) was: F =6.20 10³ L.^2.78. In females from age 2 + onwards, relative annual investment in somatic growth and reproduction was equivalent, implying that the same energy was allocated to reproduction as to maintenance/growth. Compared with other Ch. polylepis populations, the life-history patterns of this population, located in a small and seasonal stream in a southern latitude, were characterized by a low annual growth, a trade-off between reproduction and growth/maintenance, early maturity, low number of age-groups and high fecundity from multiple spawnings.     

Enzymes in cardenolide-accumulating shoot cultures of Digitalis purpurea L.

In contrast to undifferentiated cell suspension cultures of Digitalis lanata, photomixotrophic shoot cultures of Digitalis purpurea accumulate cardiac glycosides in substantial concentrations. They are used to investigate enzymes of the cardenolide pathway. All cardenolides are 5-configurated. The progesterone 5-reductase and the 3-hydroxysteroid-5-oxidoreductase are present in shoot cultures but not in undifferentiated cell cultures. These enzymes provide precursors for cardenolides, whereas the presence of the progesterone 5-reductase, also present in shoot cultures, is discussed with regard to its role in phytosterol biosynthesis and may be attributed to the general steroid pathway. The progesterone 5-reductase had an activity maximum during the early growth period seven days after onset of cultivation, whereas the corresponding progesterone 5-reductase activity was highest on day 11. The maximum cardenolide accumulation was after 24 days. The enzyme activities present in crude extracts from shoot cultures were characterized with regard to their requirements for NADPH and NADH, pH-optimum, temperature optimum, affinity to their substrates and their localization in the cell. The progesterone 5-reductase was purified 769-fold.Abbreviations DW dry weight - FW fresh weight - PVP polyvinylpyrrolidone     

A conserved region of the MSP-1 surface protein of Plasmodium falciparum contains a recognition sequence for erythrocyte spectrin.

The major surface protein MSP-1 of Plasmodium falciparum blood-stage malaria parasites contains notably conserved sequence blocks with unknown function. The recombinant protein 190L, which represents such a block, exhibits a high affinity for red blood cell membranes. We demonstrate that both 190L and native MSP-1 protein bind to the inner red blood cell membrane skeleton protein spectrin. By using overlapping peptides covering the 190L molecule, we show that the spectrin contact site of 190L is included in a linear sequence of 30 amino acid residues. Association of 190L with naturally occurring spectrin deficient red blood cells is drastically reduced. In the same cells parasite invasion is normal, but the intracellular parasite development arrests late in the trophozoite stage. A similar situation arises when synthetic peptides covering the spectrin recognition sequence of 190L are added to P.falciparum cultures. These data and the cellular localization of MSP-1 suggest the possibility that MSP-1 associates with spectrin under natural conditions.     

Selection of anthocyanin-accumulating potato (Solanum tuberosum L.) cell lines from calli derived from seedlings produced by gamma-irradiated seeds

Callus cell lines of potato (Solanum tuberosum L. cv. Zarevo) were obtained from seedlings germinated from gamma-irradiated seeds (200 Gy). Some of these cell lines produce red-violet pigments which were identified as acylated anthocyanins. The major anthocyanin was determined to be peonidin 3-O-[6-O-(4-O-E-p-coumaroyl-rhamnosyl)-glucoside]-5-O-glucoside (peonanin). Single cell-derived protoclones from non-pigmented protoplasts sometimes also gave rise to pigmented cell clusters thus indicating that the changes in the expression of the anthocyanin pathway can also occur after the stage of initial callus induction.     

Deep-water macroalgae from the Canary Islands: new records and biogeographical relationships

Due to the geographical location and paleobiogeography of the Canary Islands, the seaweed flora contains macroalgae with different distributional patterns. In this contribution, the biogeographical relations of several new records of deep-water macroalgae recently collected around the Canarian archipelago are discussed. These areBryopsidella neglecta (Berthold) Rietema,Discosporangium mesarthrocarpum (Meneghini) Hauck,Hincksia onslowensis (Amsler et Kapraun) P. C. Silva,Syringoderma floridana Henry,Peyssonnelia harveyana J. Agardh,Cryptonemia seminervis (C. Agardh) J. Agardh,Botryocladia wynnei Ballantine,Gloiocladia blomquistii (Searles) R. E. Norris,Halichrysis peltata (W. R. Taylor) P. Huvé et H. Huvé,Leptofauchea brasiliensis Joly, andSarcodiotheca divaricata W. R. Taylor. These new records, especially those in the Florideophyceae, support the strong affinity of the Canary Islands seaweed flora with the warm-temperate Mediterranean-Atlantic region. Some species are recorded for the first time from the east coast of the Atlantic Ocean, enhancing the biogeographic relations of the Canarian marine flora with that of the western Atlantic regions.     

Selection of synthetic proteins to modulate the human frataxin function

Frataxin is a kinetic activator of the mitochondrial supercomplex for iron-sulfur cluster assembly. Low frataxin expression or a decrease in its functionality results in Friedreich's Ataxia (FRDA). With the aim of creating new molecular tools to study this metabolic pathway, and ultimately, to explore new therapeutic strategies, we have investigated the possibility of obtaining small proteins exhibiting a high affinity for frataxin. In this study, we applied the ribosome display approach, using human frataxin as the target. We focused on Affi_224, one of the proteins that we were able to select after five rounds of selection. We have studied the interaction between both proteins and discussed some applications of this specific molecular tutor, concerning the modulation of the supercomplex activity. Affi_224 and frataxin showed a K_D value in the nanomolar range, as judged by surface plasmon resonance analysis. Most likely, it binds to the frataxin acidic ridge, as suggested by the analysis of chemical shift perturbations (nuclear magnetic resonance) and computational simulations. Affi_224 was able to increase Cys NFS1 desulfurase activation exerted by the FRDA frataxin variant G130V. Importantly, Affi_224 interacts with frataxin in a human cellular model. Our results suggest quaternary addition may be a new tool to modulate frataxin function in vivo. Nevertheless, more functional experiments under physiological conditions should be carried out to evaluate Affi_224 effectiveness in FRDA cell models.     

Age structure, growth and reproduction of Leuciscus pyrenaicus in an intermittent stream in the Guadalquivir river basin, southern Spain

The age, growth and reproduction of Leuciscus pyrenaicus (Günther, 1868), an endemic cyprinid from the Iberian Peninsula, was studied from November 1987 to September 1989 in a small seasonal tributary of the Guadalquivir river basin. Maximum fork lengths observed were a 160 mm male with six scale annuli and a 171 mm female aged 7 +. Maximum ages observed were 7 + in males and 8 + in females. There were no significant differences in the annual growth increments between sexes. Seasonal growth period started in March and continued for 5 to 6 months. Mean lengths of 1 + specimens onwards diminished during summer and/or autumn. Males and females matured in their third and fourth year of life respectively. The overall sex ratio (272 males: 310 females) differed significantly from equality. Spawning began in May and ended in July. L. pyrenaicus is a multiple spawner that releases a minimum of two batches of eggs per female each year. Eggs in each batch were similar in both size (egg diameter) and number released. The relationship between fecundity (Fee) and fork length (mm) was represented by the formula: Fec=1.96 10⁻³ L^2.50.