Imported disease of dogs and cats exotic to Ireland: Echinococcus multilocularis

Changes in legislation that facilitate the movement of animals within the European Union may increase the risk that some microbial and parasitic organisms, currently exotic to Ireland, will be introduced by travelled pet animals. It is possible that the fox tapeworm, Echinococcus multilocularis, might be introduced in that manner from any of the several member states in which it is endemic. Red foxes are the principal definitive hosts of E. multilocularis but dogs and cats can also be infected. Infection in the definitive host is of little clinical significance, but aberrant infection of humans results in alveolar echinococcosis, a debilitating disease that has a high mortality rate. Humans acquire the organism by ingestion of Echinococcus multilocularis eggs excreted by definitive hosts; the larval metacestodes develop primarily in the liver, in the initial asymptomatic phase as small, well-encapsulated cysts. Over time, perhaps five to 15 years, progressive local infiltration and secondary cyst development at distant sites occur with resultant clinical signs. Patients with infiltrative liver disease present with cholestatic jaundice, epigastric pain, fatigue, weight loss and hepatomegaly. If left untreated, the disease can be fatal.This paper recounts the life cycle of the parasite, and discusses the control measures on which its exclusion from Ireland depend. Strict adherence to the routine worming of travelled dogs with praziquantel, at appropriate doses, 24 to 48 hours prior to entry into the country will minimise the likelihood of introduction of this zoonosis.     

Comparison of canine cardiac troponin I concentrations as determined by 3 analyzers

BACKGROUND: Recent interest in cardiac biomarkers has led to the validation of several commercial analyzers for cardiac troponin I (cTnI) evaluation in dogs; however, these analyzers have not been standardized. HYPOTHESIS: It was hypothesized that canine plasma cTnI concentrations as determined by 3 different analyzers would be similar. ANIMALS: Twenty-three dogs with cardiac disease were studied. METHODS: Reconstituted purified canine free cTnI was diluted with canine plasma to 8 concentrations (0.01, 0.1, 0.78, 1.56, 3.13, 6.25, 12.5, and 25 ng/mL), for analysis by 3 analyzers, the Biosite Triage Meter, the Dade-Behring Stratus, and the Beckman-Coulter Access AccuTnI. Plasma samples from 23 dogs with cardiac disease were also analyzed for cTnI concentrations on all analyzers. RESULTS: Troponin I concentrations in sick dogs were <0.05-5.72 ng/mL (Biosite), 0.02-11.1 ng/mL (Access), and 0.02-9.73 ng/mL (Stratus). Analyzer results were highly correlated with each other (r = 0.97 to 1.0 for purified dilutions, r = 0.61 to 0.89 for samples from dogs); however, the limits of agreement were wide for both purified dilutions and clinical samples. Recovery was highest for the Access (334-1467%) and lowest for the Biosite (38-60%); Stratus 52-233%. Analyzer variability was lowest for the Access (1.2-10.4%) and highest for the Stratus (4.8-33.6%); Biosite 2.8-16.5%. CONCLUSIONS AND CLINICAL IMPORTANCE: Results from this study suggest that although canine cTnI values obtained from the Biosite, Stratus, and Access analyzers are closely correlated, they cannot be directly compared with each other. In the absence of a gold standard none of the analyzers can be considered more correct than the others.     

Structure of human urokinase plasminogen activator in complex with its receptor

The urokinase plasminogen activator binds to its cellular receptor with high affinity and initiates signaling cascades that are implicated in pathological processes including tumor growth, metastasis, and inflammation. We report the crystal structure at 1.9 angstroms of the urokinase receptor complexed with the urokinase amino-terminal fragment and an antibody against the receptor. The three domains of urokinase receptor form a concave shape with a central cone-shaped cavity where the urokinase fragment inserts. The structure provides insight into the flexibility of the urokinase receptor that enables its interaction with a wide variety of ligands and a basis for the design of urokinase-urokinase receptor antagonists.     

Efficient production of active recombinant Candida rugosa LIP3 lipase in Pichia pastoris and biochemical characterization of the purified enzyme

Candida rugosa lipase (CRL), an important industrial enzyme, possesses several different isoforms encoded by the high-identity lip gene family (lip1 to lip7). In this study, an additional N-terminal peptide in front of the lip3 gene was removed by PCR, and the 18 nonuniversal serine codons (CTG) of the lip3 gene were converted into universal serine codons (TCT) by means of an overlap extension PCR-based multiple-site-directed mutagenesis to express an active recombinant LIP3 in the yeast Pichia pastoris. The regional synthetic DNA fragment (339 bp) is first recombined by primer assembly with 20 overlapping nucleotides, followed by specific overlap extension PCR with outside primers containing restriction enzyme sites for directional cloning into the pGAPZalphaC vector. The results show that the production yield (0.687 unit/mL) of N-fused lip3 (nflip3) has an overall improvement of 69-fold relative to that (0.01 unit/mL) of lip3 and of 52-fold (0.47 unit/mL) of codon-optimized lip3 (colip3) relative to that (0.01 unit/mL) of non-codon-optimized lip3 (lip3), with the cultivation time set at 5 days. This finding demonstrates that the reservation of the N terminus and the regional codon optimization of the lip3 gene fragment at the 5' end can greatly increase the expression level of recombinant LIP3 in the P. pastoris system. The purified recombinant LIP3 shows distinct biochemical properties compared with other isoforms.     

Candida rugosa lipase LIP1-catalyzed transesterification to produce human milk fat substitute

Structured lipids (SLs) containing palmitic and oleic acids were synthesized by transesterification of tripalmitin with either oleic acid or methyl oleate as acyl donor. This SL with palmitic acid at the sn-2 position and oleic acid at sn-1,3 positions is similar in structure to human milk fat triacylglycerol. LIP1, an isoform of Candida rugosa lipase (CRL), was used as biocatalyst. The effects of reaction temperature, substrate molar ratio, and time on incorporation of oleic acid were investigated. Reaction time and temperature were set at 6, 12, and 24 h, and 35, 45, and 55 degrees C, respectively. Substrate molar ratio was varied from 1:1 to 1:4. The highest incorporation of oleic acid (37.7%) was at 45 degrees C with methyl oleate as acyl donor. Oleic acid resulted in slightly lesser (26.3%) incorporation. Generally, higher percentage incorporation of oleic acid was observed with methyl oleate (transesterification) than with oleic acid (acidolysis). In both cases percentage incorporation increased with reaction time. Incorporation decreased with increase in temperature above 45 degrees C. Initially, oleic acid incorporation increased with increase in substrate molar ratio up to 1:3. LIP1 was also compared with Lipozyme RM IM as biocatalysts. The tested reaction parameters were selected on the basis of maximum incorporation of C18:1 obtained during optimization of LIP1 reaction conditions. Reaction temperature was maintained at 45, 55, and 65 degrees C. Lipozyme RM IM gave highest oleic acid incorporation (49.4%) at 65 degrees C with methyl oleate as acyl donor. Statistically significant (P < 0.05) differences were observed for both enzymes. SL prepared using Lipozyme RM IM may be more suitable for possible use in human milk fat substitutes.     

Waking experience affects sleep need in Drosophila

Sleep is a vital, evolutionarily conserved phenomenon, whose function is unclear. Although mounting evidence supports a role for sleep in the consolidation of memories, until now, a molecular connection between sleep, plasticity, and memory formation has been difficult to demonstrate. We establish Drosophila as a model to investigate this relation and demonstrate that the intensity and/or complexity of prior social experience stably modifies sleep need and architecture. Furthermore, this experience-dependent plasticity in sleep need is subserved by the dopaminergic and adenosine 3',5'-monophosphate signaling pathways and a particular subset of 17 long-term memory genes.