Uncommon and fatal case of cystoisosporiasis in a non HIV-immunosuppressed patient from a non-endemic country

Cystoisospora belli (previously known as Isospora belli) is a tropical coccidian parasite sometimes leading to severe diarrhea in immunocompromised patients. Here we describe a fatal case of cystoisosporiasis in a non HIV-immunocompromised 71-year-old female with no recent travel history. Infection was either latent or potentially caused by the consumption of contaminated imported food from Asia. Diagnosis was made by microscopical detection of numerous C. belli oocysts in stools without specific staining. Treatment with TMP-SMZ slightly improved diarrhea within 3 days, but dehydration subsequently led to acute decompensated heart failure and a fatal evolution. This report illustrates the possibility of severe cystoisosporiasis in non HIV-immunocompromised patients in a non-endemic country and highlights the risk of transmission through imported contaminated food consumption.     

Isolation,characterization and molecular cloning of the mannose-binding lectins from leaves and roots of garlic (Allium sativum L.)

Two novel lectins were isolated from roots and leaves of garlic. Characterization of the purified proteins indicated that the leaf lectin ASAL is a dimer of two identical subunits of 12 kDa, which closely resembles the leaf lectins from onion, leek and shallot with respect to its molecular structure and agglutination activity. In contrast, the root lectin ASARI, which is a dimer of subunits of 15 kDa, strongly differs from the leaf lectin with respect to its agglutination activity. cDNA cloning of the leaf and root lectins revealed that the deduced amino acid sequences of ASAL and ASARI are virtually identical. Since both lectins have identical N-terminal sequences the larger Mr of the ASARI subunits implies that the root lectin has an extra sequence at its C-terminus. These results not only demonstrate that virtually identical precursor polypeptides are differently processed at their C-terminus in roots and leaves but also indicate that differential processing yields mature lectins with strongly different biological activities. Further screening of the cDNA library for garlic roots also yielded a cDNA clone encoding a protein composed of two tandemly arrayed lectin domains. Since the presumed two-domain root lectin has not been isolated yet, its possible relationship to the previously described two-domain bulb lectin could not be studied at the protein level.     

Reconstituted human gingival epithelium: Nonsubmerged in vitro model

Summary Many studies have shown that human gingival keratinocytes grown in submerged culture fail to attain optimal differentiation. This study reports an in vitro culture system for oral gingival epithelial cells, in which they are grown at the air-liquid interface, on polycarbonate inserts, in the presence of an NIH-3T3 feeder layer. This model was compared with two submerged culture methods for gingival keratinocytes, on type I collagen gel and on an NIH-3T3 feeder layer. Transmission electron microscopy showed an advanced level of stratification (over six layers of cells) for cultures grown at the air-liquid interface. Immunofluorescence and electrophoretic patterns showed the presence of cytokeratins 10 and 11 in cytoskeletal protein extracts of these cultured keratinocytes. In this air-liquid interface culture model, in the presence of NIH-3T3 feeder cells, keratinocytes can achieve an advanced level of stratification and differentiation and a resemblance to in vivo gingiva. The obtention of a highly differentiated epithelium will permit in vitro pharmacological studies and studies on the biocompatability of certain alloys with the superficial periodontium; it will also provide grafts for patients undergoing periodontal surgery.     

MORPHOLOGICAL AND MOLECULAR CHARACTERIZATION OF PLANKTONIC CYANOBACTERIA FROM BELGIUM AND LUXEMBOURG1

For the first time in Belgium and Luxembourg, the diversity and taxonomy of 95 cyanobacterial strains isolated from freshwater blooms were assessed by the comparison of phenotypes and partial 16S rRNA gene sequences. The results showed the high diversity of nanoplanktonic, picoplanktonic, and benthic–periphytic cyanobacteria accompanying the main bloom‐forming taxa. Indeed, besides 15 morphotypes of bloom‐forming taxa, seven non‐bloom‐forming planktonic morphotypes and 11 morphotypes from benthic–periphytic taxa were isolated in culture from the plankton samples of 35 water bodies. The bloom‐forming strains belonged to the genera Microcystis, Woronichinia, Planktothrix, Anabaena, and Aphanizomenon, whereas the other strains isolated from the same samples were assigned to the nanoplanktonic Aphanocapsa, Aphanothece, Snowella, and Pseudanabaena; to the picoplanktonic Cyanobium; and to the benthic–periphytic Geitlerinema, Komvophoron, Leptolyngbya, Lyngbya, Phormidium, Calothrix, Nostoc, and Trichormus. The results supported both the polyphyletism of genera such as Aphanocapsa, Aphanothece, Leptolyngbya, Geitlerinema, Anabaena, and Aphanizomenon as well as the validity of genera such as Microcystis, Planktothrix, and Pseudanabaena with gas vesicles and cells constricted at the cross wall. The results obtained showed the close relationship between Snowella and Woronichinia for which very few sequences exist. The first sequence of Komvophoron appeared poorly related to other available cyanobacterial sequences. Although in a few cases a good agreement existed between phenotypic and genotypic features, there was generally a discrepancy. Strains with identical morphotypes show small differences in the 16S rRNA sequences, which might be related to the different chemical properties of their habitats. The results showed the importance of the polyphasic approach in order to improve the taxonomy of cyanobacteria.     

Gastric HCO3-stimulated ATPase: Evidence against its microsomal localization in rat fundus mucosa

Rat gastric mucosa was shown to contain a Mg²⁺-dependent ATPase which is stimulated by HCO₃⁻ at pH 8–9.Triton X-100 solubilizes this HCO₃⁻-stimulated, Mg²⁺-dependent ATPase (ATP phosphohydrolase, EC 3.6.1.3).The gastric mucosa was resolved into five subcellular fractions by differential centrifugation. A large granule fraction (Fraction M), 28 000 g · min, was characterized by cytochrome c oxidase (marker enzyme for mitochondria). A microsomal fraction (Fraction P), 2 760 000 g · min, was characterized by 5′-nucleotidase(5′-ribonucleotide phosphohydrolase, EC 3.1.3.5) (plasma membrane).The Mg²⁺-dependent ATPase was demonstrated to have a bimodal mitochondrial membranous localization: 24% of its activity is associated with cytochrome c oxidase, and 75% with 5′-nucleotidase(5′-ribonucleotide phosphohydrolase, EC 3.1.3.5) at pH 8.The HCO₃⁻ addition resulted in two opposite effects: (1) a strong stimulation (84%) in Fraction M; (2) a slight inhibition (12%) in Fraction P.Fraction M was subfractionated by equilibration on a sucrose gradient. It gave rise to a homogeneous mitochondrial (d, 1.17–1.21) Mg²⁺-dependent ATPase, closely associated with cytochrome c oxidase. This ATPase is strongly stimulated (×2) by HCO₃⁻. The subfractionation of Fraction P gave rise to two distinct ATPases: (1) the major one is associated with membranous (d, 1.10–1.15) material marked by 5′-nucleotidase and is slightly inhibited by HCO₃⁻; (2) the other is associated with denser (d, 1.17–1.21) material and is stimulated by HCO₃⁻.The bicarbonate-stimulated fraction of the Mg²⁺-dependent ATPase activity found in the gastric microsomal fraction is assumed to arise from mitochondrial cross-contamination. Further support comes from the optimal HCO₃⁻ concentration. In addition, SCN⁻ is shown to specifically inhibit the ATPase of Fraction M.From these results it appears that the implication of HCO₃⁻-stimulated ATPase in the gastric secretion of H⁺ is not as clear as had been suggested. However, in the view of an ATPase-supported model for H⁺ secretion, attention can be directed towards the Mg²⁺-dependent ATPase found to be associated with microsomes.     

The structural and functional organization of H-NS-like proteins is evolutionarily conserved in Gram-negative bacteria

The structural gene of the H-NS protein, a global regulator of bacterial metabolism, has been identified in the group of enterobacteria as well as in closely related bacteria, such as Erwinia chrysanthemi and Haemophilus influenzae . Isolated outside these groups, the BpH3 protein of Bordetella pertussis exhibits a low amino acid conservation with H-NS, particularly in the N-terminal domain. To obtain information on the structure, function and/or evolution of H-NS, we searched for other H-NS-related proteins in the latest databases. We found that HvrA, a trans -activator protein in Rhodobacter capsulatus , has a low but significant similarity with H-NS and H-NS-like proteins. This Gram-negative bacterium is phylogenetically distant from Escherichia coli . Using theoretical analysis (e.g. secondary structure prediction and DNA binding domain modelling) of the amino acid sequence of H-NS, StpA (an H-NS-like protein in E. coli ), BpH3 and HvrA and by in vivo and in vitro experiments (e.g. complementation of various H-NS-related phenotypes and competitive gel shift assay), we present evidence that these proteins belong to the same class of DNA binding proteins. In silico analysis suggests that this family also includes SPB in R. sphaeroides , XrvA in Xanthomonas oryzae and VicH in Vibrio cholerae . These results demonstrate that proteins structurally and functionally related to H-NS are widespread in Gram-negative bacteria.     

Molecular Determinants of the Interaction Between the C-Terminal Domain of Alzheimer's β-Amyloid Peptide and Apolipoprotein E α-Helices

In a previous work, we predicted and demonstrated that the 29-42-residue fragment of beta-amyloid peptide (Abeta peptide) has in vitro capacities close to those of the tilted fragment of viral fusion proteins. We further demonstrated that apolipoprotein E2 and E3 but not apolipoprotein E4 can decrease the fusogenic activity of Abeta(29-42) via a direct interaction. Therefore, we suggested that this fragment is implicated in the neurotoxicity of Abeta and in the protective effects of apolipoprotein E in Alzheimer's disease. Because structurally related apolipoproteins do not interact with the Abeta C-terminal domain but inhibit viral fusion, we suggested that interactions existing between fusogenic peptides and apolipoproteins are selective and responsible for the inhibition of fusion. In this study, we simulated interactions of all amphipathic helices of apolipoproteins E and A-I with Abeta and simian immunodeficiency virus (SIV) fusogenic fragments by molecular modeling. We further calculated cross-interactions that do not inhibit fusion in vitro. The results suggest that interactions of hydrophobic residues are the major event to inhibit the fusogenic capacities of Abeta(29-42) and SIV peptides. Selectivity of those interactions is due to the steric complementarity between bulky hydrophobic residues in the fusogenic fragments and hydrophobic residues in the apolipoprotein C-terminal amphipathic helices.