Molecularly Imprinted Polymers for the isolation of bioactive naphthoquinones from plant extracts

Molecularly Imprinted Polymers (MIPs) targeting shikonin, a potent antioxidant and wound healing agent, have been prepared using methacrylic acid (MAA) and 2-diethylaminoethyl methacrylate (DEAEMA) as functional monomers. An investigation of solution association between shikonin and both acidic and basic functional monomers by UV-Vis titrations, suggested stronger affinity towards the basic functionality. Strong inhibition of the co-polymerisation reaction of such basic monomers was observed, but was overcome by reduction of the amount of template used during polymer synthesis. Polymer morphology was severely impacted by the template’s radical scavenging behaviour as demonstrated by solid state NMR spectroscopy measurements. HPLC evaluation of the final materials in polar conditions revealed limited imprinting effects and selectivity, with the MAA polymers exhibiting marginally better performance. During application of the polymers as MI-SPE sorbents in non-polar solvents it was found that the DEAEMA based polymer was more selective towards shikonin compared to the MAA counterpart, while shikonin recoveries of up to 72% were achieved from hexane solutions of a commercial sample of shikonin, hexane extract of Alkanna tinctoria roots and a commercial pharmaceutical ointment.

Habitat Isolation Reduces the Temporal Stability of Island Ecosystems in the Face of Flood Disturbance

Natural ecosystems are increasingly exposed to multiple anthropogenic stressors, including land-use change, deforestation, agricultural intensification, and urbanisation, all of which have led to widespread habitat fragmentation, which is also likely to be amplified further by predicted climate change. The potential interactive effects of these different stressors cannot be determined by studying each in isolation, although such synergies have been largely ignored in ecological field studies to date. Here, we use a model system of naturally fragmented islands in a braided river network, which is exposed to periodic inundation, to investigate the interactive effects of habitat isolation and flood disturbance. Food web structure was similar across the islands during periods of hydrological stability, but several key properties were altered in the aftermath of flood disturbance, based on distance of the islands from the regional source pool of species: taxon richness and mean food chain length declined with habitat isolation after flooding, while the proportion of basal species increased. Greater species turnover through time reflected the slower process of re-colonisation on the more distant islands following disturbance. Increased variability of several food web properties over a 1-year period highlighted the reduced temporal stability of isolated habitat fragments. Many of these effects reflected the differential successes of predator and prey species at re-colonising the islands: even though larger, more mobile consumers may reach the more distant islands first, they cannot establish populations until the lower trophic levels have successfully reassembled. These results highlight the susceptibility of fragmented ecosystems to environmental perturbations. © 2013 Elsevier Ltd.

Isolation of promoter for cytosolic phospholipase A2 (cPLA2)

Cytosolic phospholipase A2 (cPLA2) releases arachidonic acid from membrane phospholipids and is believed to be the rate-limiting enzyme in the arachidonic acid pathway. We report herein the isolation of a 3 kb fragment of rodent genomic DNA containing part of the first intron, the first exon and 5'-flanking sequence. The start site of transcription was mapped by 5'-rapid amplification of cDNA ends and corroborated by ribonuclease protection assay. The gene has a TATAless promoter with no classical Sp1 binding sites or initiator element. A microsatellite series of CA repeats was noted in the 5'-flanking region of both the rodent and human promoters. Deletion constructs have been analysed for luciferase activity and confirmed promoter activity.

Isolation and characterization of eight polymorphic microsatellite loci for Natterer's bat, Myotis nattereri (Vespertilionidae, Chiroptera)

Eight new microsatellite loci were isolated and characterized for the Natterer's bat Myotis nattereri from a microsatellite-enriched genomic library. The usefulness of these markers was assessed by screening a sample comprising 100 specimens collected from throughout the species range in Europe. Both moderately and highly polymorphic loci were identified with 3-17 alleles segregating per locus (mean 8.1 SE +/- A 0.048). No evidence for departure from HWE or linkage disequilibrium among loci was observed. These markers will provide a valuable addition to the molecular toolbox currently available for studies of population genetic structure, parentage and social organisation of M. nattereri and related species.

Rifampicin and sodium fusidate reduces the frequency of methicillin-resistant Staphylococcus aureus (MRSA) isolation in adults with cystic fibrosis and chronic MRSA infection

Nosocomial transmission of methicillin-resistant Staphylococcus aureus (MRSA) to patients with cystic fibrosis (CF) frequently results in chronic respiratory tract carriage. This is an increasing problem, adds to the burden of glycopeptide antibiotic use in hospitals, and represents a relative contraindication to lung transplantation. The aim of this study was to determine whether it is possible to eradicate MRSA with prolonged oral combination antibiotics, and whether this treatment is associated with improved clinical status. Adult CF patients (six male, one female) with chronic MRSA infection were treated for six months with rifampicin and sodium fusidate. Outcome data were examined for six months before treatment, on treatment and after treatment. The patients had a mean age of 29.3 (standard deviation=6.3) years and FEV(1) of 36.1% (standard deviation=12.7) predicted. The mean duration of MRSA isolation was 31 months. MRSA isolates identified in these patients was of the same lineage as the known endemic strain at the hospital when assessed by pulsed-field gel electrophoresis. Five of the seven had no evidence of MRSA during and for at least six months after rifampicin and sodium fusidate. The proportion of sputum samples positive for MRSA was lower during the six months of treatment (0.13) and after treatment (0.19) compared with before treatment (0.85) (P<0.0001). There was a reduction in the number of days of intravenous antibiotics per six months with 20.3+/-17.6 on treatment compared with 50.7 before treatment and 33.0 after treatment (P=0.02). There was no change in lung function. Gastrointestinal side effects occurred in three, but led to therapy cessation in only one patient. Despite the use of antibiotics with anti-staphylococcal activity for treatment of respiratory exacerbation, MRSA infection persists. MRSA can be eradicated from the sputum of patients with CF and chronic MRSA carriage by using rifampicin and sodium fusidate for six months. This finding was associated with a significant reduction in the duration of intravenous antibiotic treatment during therapy.

An optimised milk testing protocol to ensure accurate enumeration of viable Mycobacterium avium subsp. paratuberculosis by the PMS-phage assay

There is interest in determining levels of Mycobacterium avium subsp. paratuberculosis (MAP) contamination in milk. The optimal sample preparation for raw cows' milk to ensure accurate enumeration of viable MAP by the peptide-mediated magnetic separation (PMS)-phage assay was determined. Results indicated that milk samples should be refrigerated at 4 C after collection and MAP testing should commence within 24 h, or samples can be frozen at 70 C for up to one month without loss of MAP viability. Use of Bronopol is not advised as MAP viability is affected. The vast majority (>95%) of MAP in raw milk sedimented to the pellet upon centrifugation at 2500 g for 15 min, so this milk fraction should be tested. De-clumping of MAP cells was most effectively achieved by ultrasonication of the resuspended milk pellet on ice in a sonicator bath at 37 kHz for 4 min in ‘Pulse’ mode.