Hepatosplenic candidiasis, a late manifestation of Candida septicaemia in neutropenic patients with haematologic malignancies

Systemic candidiasis with Candida-induced abscesses, predominantly in the liver and the spleen, was diagnosed in 27 patients with haematologic malignancies after intensive cytostatic therapy. Specific features included septic fever unresponsive to antimicrobial therapy, hepatosplenomegaly with multiple lesions in the liver and spleen (diameter up to 2 cm) as detected by computed tomography (CT) or ultrasound, and an elevation in liver enzymes. During treatment, induced neutropenia, hepatic and splenic foci were poorly defined histologically and were not identified by imaging procedures. After granulocyte recovery these foci showed characteristic histological patterns. Ultrasound and/or CT investigations of the abdomen now revealed characteristic lesions in the liver and the spleen. Gamma-GT and alkaline phosphatase were early indicators of hepatic involvement in Candida septicaemia and were often already elevated in aplasia.     

Ca2+ and cAMP activate K+ channels in the basolateral membrane of crypt cells isolated from rabbit distal colon

Summary Using patch-clamp techniques, we have studied Ca²⁺-activated K⁺ channels in the basolateral membrane of freshly isolated epithelial cells from rabbit distal colon. Epithelial cell clusters were obtained from distal colon by gentle mechanical disruption of isolated crypts. Gigaohm seals were obtained on the basolateral surface of the cell clusters. At the resting potential (approximately –45 mV), with NaCl Ringer's bathing the cell, the predominant channels had a conductance of 131±25 pS. Channel activity depended on voltage as depolarization of the membrane increased the open probability. In excised inside-out patches, channels were found to be selective for K⁺ over Na⁺. Channel activity correlated directly with bath Ca²⁺ concentration in the excised patches. Channel currents were blocked by 5mm TEA⁺ and 1mm Ba²⁺. In cell-attached patches, after addition of the Ca²⁺ ionophore A23187, which increases intracellular Ca²⁺, open probability was markedly increased. Channel activity was also regulated by cAMP as addition of 1mm dibutyryl-cAMP in the bath solution in cell-attached patches increased channel open probability over 20-fold. Channels that had been activated by cAMP were further activated by Ca²⁺. We conclude that the basolateral membrane of epithelial cells from descending colon contains a class of potassium channels, which are regulated by intracellular Ca²⁺ and cAMP.     

Trimethoprim resistance transposon Tn4003 from Staphylococcus aureus encodes genes for a dihydrofolate reductase and thymidylate synthetase flanked by three copies of IS257

Trimethoprim resistance mediated by the Staphylococcus aureus multi-resistance plasmid pSK1 is encoded by a structure with characteristics of a composite transposon which we have designated Tn4003. Nucleotide sequence analysis of Tn4003 revealed it to be 4717 bp in length and to contain three copies of the insertion element IS257 (789-790 bp), the outside two of which are flanked by directly repeated 8-bp target sequences. IS257 has imperfect terminal inverted repeats of 27-28 bp and encodes for a putative transposase with two potential alpha-helix-turn-alpha-helix DNA recognition motifs. IS257 shares sequence similarities with members of the IS15 family of insertion sequences from Gram-negative bacteria and with ISS1 from Streptococcus lactis. The central region of the transposon contains the dfrA gene that specifies the S1 dihydrofolate reductase (DHFR) responsible for trimethoprim resistance. The S1 enzyme shows sequence homology with type I and V trimethoprim-resistant DHFRs from Gram-negative bacteria and with chromosomally encoded DHFRs from Gram-positive and Gram-negative bacteria. 5' to dfrA is a thymidylate synthetase gene, designated thyE.     

Secondary Invasions: Implications of Riparian Restoration for In-Stream Invasion by an Aquatic Grass

The complex nature of ecosystems with multiple invaders requires whole-system approaches to ecosystem management. Undesirable, unintended secondary effects may occur if interspecific interactions are ignored. Although degraded riparian zones urgently need effective restoration, broad-scale removal of introduced tree species (e.g., willows [ Salix spp.]) and fencing of riparian zones to exclude livestock may facilitate spread of the invasive aquatic grass Glyceria maxima in southeastern Australia. We recorded occurrence of the grass at riparian sites with different amounts of woody vegetation, including willows, and monitored spread for 2 years in locations with and without livestock. Glyceria maxima occurred more frequently at sites with little or no woody riparian vegetation. Larger, older patches fenced from livestock spread fastest. Analyses of costs of controlling G. maxima with herbicide showed that it is more cost-effective to eradicate small patches than large patches if locations are known. However, the cost per m² reduction in patch size is cheaper for larger patches. We recommend that small, young patches should be eradicated as soon as detected and show that spread of large source patches can be controlled effectively with continued spraying over several years. If restoration of waterways is to succeed, riparian management strategies must recognize connectivity between riparian and freshwater habitats.     

Freshwater invasions: using historical data to analyse spread

Aquatic invasive species cause deleterious environmental and economic impacts, and are rapidly spreading through ecosystems worldwide. Despite this, very few data sets exist that describe both the presence and the absence of invaders over long time periods. We have used Geographical Information Systems (GIS) to analyse time-series data describing the spread of the freshwater invasive New Zealand mudsnail, Potamopyrgus antipodarum , in Victoria, Australia, over 110 years. We have mapped the snail's spread, estimated the percentage of stream length invaded through time, calculated the functional form of the spread rate, and investigated the role that the two proposed vectors — fish stocking and angling — have had in this invasion. Since it was first found in 1895, P. antipodarum has expanded its range in Victoria and now occurs throughout much of the southern and central areas of the state. The north of the state is relatively less invaded than the south, with the division corresponding approximately to the presence of the Great Dividing Range. We show that the snail's range has been increasing at an approximately exponential rate and estimate that 20% of total Victorian stream length is currently invaded. We also show that using long-term data can change the outcome of analyses of the relationship between vectors of spread and invasion status of separate catchments. When our time-series data were aggregated through time, the total numbers of fish stocking events and angling activity were both correlated with invasion. However, when the time-series data were used and the number of fish stocking events calculated up until the date of invasion, no relationships with stocking were found. These results underline the role that time-series data, based on both presences and absences, have to play when investigating the spread of invasive species.     

Male-Specific Association between a γ-Secretase Polymorphism and Premature Coronary Atherosclerosis

Background

Atherosclerosis is a common multifactorial disease resulting from an interaction between susceptibility genes and environmental factors. The causative genes that contribute to atherosclerosis are elusive. Based on recent findings with a Wistar rat model, we speculated that the γ-secretase pathway may be associated with atherosclerosis.

Methodology/Principal Findings

We have tested for association of premature coronary atherosclerosis with a non-synonymous single-nucleotide polymorphism (SNP) in the γ-secretase component APH1B (Phe217Leu; rs1047552), a SNP previously linked to Alzheimer''s disease. Analysis of a Dutch Caucasian cohort (780 cases; 1414 controls) showed a higher prevalence of the risk allele in the patients (odds ratio (OR) = 1.35), albeit not statistically different from the control population. Intriguingly, after gender stratification, the difference was significant in males (OR = 1.63; p = 0.033), but not in females (OR = 0.50; p = 0.20). Since Phe217Leu-mutated APH1B showed reduced γ-secretase activity in mouse embryonic fibroblasts, the genetic variation is likely functional.

Conclusion/Significance

We conclude that, in a male-specific manner, disturbed γ-secretase signalling may play a role in the susceptibility for premature coronary atherosclerosis.     

Identifying interacting residues using Boolean Learning and Support Vector Machines: case study on mRFP and DsRed proteins

In a protein, interactions exist between amino acid residues that influence the protein's structural integrity or stability and thus affect its catalytic function. The loss of this interaction due to mutations in these amino acids usually leads to a non-functional protein. Probing the sequence space of a protein through mutations or recombinations, as performed in directed evolution to search for an improved variant, frequently results in such inactive sequences. In this work, we demonstrate the use of machine learning to identify such interacting residues and the use of template engineering strategies to increase the fraction of active variants in a library. We show that using the sequences from recombination of monomeric red fluorescent protein (mRFP) and Discosoma red fluorescent protein (DsRed), we were able to identify a pair of interacting residues using an algorithm based on Boolean Learning and Support Vector Machines. The interaction between the identified residues was verified through point mutations on the mRFP and DsRed genes. We also show that it is possible to use such results to alter the parental genes such that the probability of disrupting the important interactions is minimized. This will result in a larger fraction of active variants in the recombinant library and allow us to access more functional space. We demonstrate this effect by comparing the recombinant library of wild-type (WT) DsRed, mRFP and an altered sequence of DsRed with mRFP WT genes.     

Ataxin-2 associates with the endocytosis complex and affects EGF receptor trafficking

Ataxin-2 is a novel protein, where the unstable expansion of an internal polyglutamine domain can cause the neurodegenerative disease Spinocerebellar Ataxia type 2 (SCA2). To elucidate its cellular function, we have used full-length ataxin-2 as bait in a yeast two-hybrid screen of human adult brain cDNA. As binding partners we found endophilin A1 and A3, two brain-expressed members of the endophilin A family involved in synaptic vesicle endocytosis. Co-immunoprecipitation studies confirmed the binding of these proteins as an endogenous complex in mouse brain. In vitro binding experiments narrowed the binding interfaces down to two proline-rich domains on ataxin-2, which interacted with the SH3 domain of endophilin A1/A3. Ataxin-2 and endophilin associated at the endoplasmic reticulum as well as at the plasma membrane as determined by immunofluorescence microscopy of transfected cell lines, and by centrifugation fractionation studies of mouse brain. Importantly, the pattern observed in transfected cells was conserved in rat hippocampal neurons. In the mouse brain, an association of ataxin-2 with endocytic proteins such as the adaptor CIN85 and the ubiquitin ligase c-Cbl was also demonstrated. GST pull-down assays showed ataxin-2 to directly interact with the SH3 domains A and C of CIN85 and with the SH3 domain of Src, a kinase activated after receptor stimulation. Functional studies demonstrated that ataxin-2 affects endocytic trafficking of the epidermal growth factor receptor (EGFR). Taken together, these data implicate ataxin-2 to play a role in endocytic receptor cycling.