Neurocognitive function in procedures correcting severe aortic valve stenosis: patterns and determinants.

Background: Neurocognitive changes occurring after a surgical aortic valve replacement (SAVR) or transcatheter aortic valve implantation (TAVI) procedure for the correction of severe aortic stenosis (AS) have not been widely addressed and, if addressed, have produced conflicting results. The purpose of this study is to identify the pre-procedural neurocognitive pattern and its determinants in a setting of elderly (>65 years) patients with severe AS undergoing SAVR or TAVI and the changes occurring at a 2-3 month follow-up. Methods: This was a prospective cohort study included in the Italian Registry on Outcomes in Aortic Stenosis Treatment in Elderly Patients. Patients were assessed both before and after (2-3 months) the procedure using the Montreal Cognitive Assessment (MoCA) test. Data on periprocedural demographics, clinical factors, and outcome measures were collected. Results: Before the procedure, 70% of the patients demonstrated a MoCA score <23 points, which was indicative of cognitive dysfunction. The factors associated with neurocognitive dysfunction were age, functional capacity, chronic heart failure, and hemoglobin levels. After the procedure, there was an overall improvement in the MoCA score of the patients, but 28% of the patients showed a reliable worsening of their condition. The factors associated with MoCA worsening were platelet transfusions and the amount of red blood cell units transfused. Conclusion: The correction of severe AS leads to an improvement in neurocognitive function after 2-3 months. This improvement does not differentiate between SAVR and TAVI after matching for pre-procedural factors. The only modifiable factor associated with pre-procedural neurocognitive function is anemia, and anemia correction with red blood cell transfusions is associated with a worsening of neurocognitive function. This leads to the hypothesis that anemia correction before the procedure (with iron and/or erythropoietin) may limit the risk of a post-procedural worsening of neurocognitive function.

A method for collecting large quantities of Cryptosporidium parasites.

The intestinal mucoso may be regarded as a potential and abundant source of Cryptosporidium parvurn parasites from which all developmental stages might be collected. If intracellular stages could be recovered from the brush border, many of the limitations concerned with the use of oocysts and in vitro cultures may be overcome. Hans-Michael Muller, Lorella Ranucci, Edoordo Pozio and Andrea Crisonti discuss here how this can be done.

Organization of aerobactin, hemolysin, and antibacterial resistance genes in lactose-negative Escherichia coli strains of serotype O4 isolated from children with diarrhea.

Epidemiologically related, non-lactose-fermenting (NLF) Escherichia coli strains of serotype O4 have been isolated at a high frequency from children with diarrhea in Somalia (M. Nicoletti, F. Superti, C. Conti, A. Calconi, and C. Zagaglia, J. Clin. Microbiol. 26:524-529, 1988). In order to define the virulence potential of these strains, we characterized the replication properties of their high-molecular-weight plasmids and studied the genetic locations and organization of the aerobactin (aer) and hemolysin (hly) determinants encoded by 23 NLF O4 E. coli strains. Southern blot hybridizations, mobilization assays of nonconjugative plasmids, and incompatibility-exclusion experiments conducted with a conjugative incompatibility group FI (IncFI) plasmid showed that (i) 20 out of the 23 strains examined harbor a 160- to 180-kb IncFI plasmid that shares homology with the basic replicons RepFIA, RepFIB, and (except for the plasmid of one strain) RepFIC, and 22 strains also contain a 40- to 140-kb IncFII plasmid sharing homology with the RepFIIA replicon; (ii) the IncFI plasmid is nonconjugative and carries antibiotic resistance genes; (iii) the aer system is located on the IncFI plasmids and/or the chromosomes in the three strains not harboring IncFI, and it is found in an inverted orientation; (iv) the hly determinants are located on the chromosome, and their genetic organization is well conserved and closely resembles that of the reference hemolytic plasmid pHly152; and (v) Hly- mutants obtained by transposon insertion mutagenesis are not cytotoxic to HeLa cell monolayers, indicating that hemolysin is responsible for the high cytotoxic activity we have previously reported for these strains. The structural organization of the plasmid-encoded aer operon, together with the finding that those plasmids also carry antibiotic resistance genes, indicates that the IncFI plasmid of the NLF O4 E. coli strains studied more closely resembles aer-encoding virulence IncFI Salmonella R plasmids than E. coli ColV plasmids. The data presented here cannot rule out whether the strains examined are potentially intestinal or extraintestinal pathogens. Nevertheless, the genetic organization of the virulence genes, together with the epidemiological behavior and the wide spectrum of antibiotic resistance of the NLF O4 E. coli strains, indicates that these strains are structured as typical E. coli pathogenic isolates of human origin.

Cloning of the entire COWP gene of Cryptosporidium parvum and ultrastructural localization of the protein during sexual parasite development.

Molecular cloning and immunoelectron microscopy have been used to clone the full-length gene encoding Cryptosporidium parvum oocyst wall protein (COWP) and to analyse at the ultrastructural level the expression and localization of COWP during development in the gut. COWP is 1622 amino acids long, has a typical leader peptide and consists of 2 amino acidic domains each containing distinct repeated elements possibly originating from a common ancestral precursor. Electron microscopy localized COWP in a large cytoplasmic inclusion and in the wall-forming bodies of early and late macrogametes, respectively. Ultrastructural analysis of double-walled sporulating and mature oocysts indicated that COWP is selectively localized in the inner layer of the oocyst wall. This study provides the first localization at the ultrastructural level of a cloned coccidian oocyst wall protein.

Molecular cloning and expression of a 70-kilodalton heat shock protein of Candida albicans.

By screening an expression library of the yeast form of Candida albicans with a serum directed against whole fungal cells, a cDNA (2,325 bp) encoding a stress protein of C. albicans was cloned and sequenced. The cloned sequence (CaRLV130) identified a single open reading frame with a length of 1,968 bp coding for a protein containing 656 amino acid residues (70 kDa). The deduced amino acid sequence was 84% similar to the sequence of the Saccharomyces cerevisiae SSA1 gene, which encodes one member of the 70-kDa heat shock protein (Hsp70) family. The relevant gene (C. albicans HSP70 gene [CaHSP70]) was localized on the highest-M(r) (R1; approximately 3.8 Mb) chromosome of C. albicans as determined by pulse-field electrophoresis. CaHSP70 was expressed after heat shock, as demonstrated by Northern (RNA) blotting and reverse transcriptase-PCR with specific pairs of oligonucleotide sequences and gene probes. A recombinant protein was obtained in Escherichia coli after cloning of the full coding sequence into the BamHI site of the pDS56/RBSII6xhisE- plasmid and purification by nickel chelate affinity chromatography. The recombinant protein (6xhis-CaHsp70) was efficiently recognized in immunoblots by a monoclonal antibody directed against a common epitope of eukaryotic Hsp70 proteins, as well as by sera from normal human subjects. Moreover, immune mouse sera against the purified recombinant protein recognized native, heat-inducible constituents with sizes of around 70 kDa in whole-cell protein extracts of C. albicans. Overall, our data demonstrate that CaHSP70 encodes one member of a family of proteins (Hsp70) which usually represent highly conserved immunodominant antigens of infectious agents.

Characterization and immunolocalization of a Cryptosporidium protein containing repeated amino acid motifs.

The oocyst wall is one of the components that permits cryptosporidia both to survive in the environment and to retain infectivity. With the aim of identifying Cryptosporidium proteins specifically expressed at the oocyst stage, we screened lambda gt11 genomic libraries of Cryptosporidium parvum with both an oocyst antiserum and a specific genetic probe. We isolated, from distinct libraries, two overlapping clones containing an open reading frame encoding a 1,252-amino-acid polypeptide. The analysis of the deduced amino acid sequence revealed unusually high contents of cysteine, proline, and histidine. The sequence was also characterized by two distinct amino acid motifs, each repeated several times. The DNA sequences coding for the amino acid repeats showed a high frequency of synonymous mutations, a result suggesting that the repeated motifs may be functionally and/or structurally important to the parasite. Antisera and monoclonal antibodies developed against a recombinant polypeptide encompassing the first 786 amino acids revealed that the corresponding protein in C. parvum had an apparent molecular weight of 190,000. Moreover, confocal microscopy analysis with immunofluorescence indicated that the protein was localized on the oocyst wall as a uniform stain and within the oocyst itself as bright granules in close association with the residual body.