Nitrous Oxide Reductase (nosZ) Gene Fragments Differ between Native and Cultivated Michigan Soils

The effect of standard agricultural management on the genetic heterogeneity of nitrous oxide reductase (nosZ) fragments from denitrifying prokaryotes in native and cultivated soil was explored. Thirty-six soil cores were composited from each of the two soil management conditions. nosZ gene fragments were amplified from triplicate samples, and PCR products were cloned and screened by restriction fragment length polymorphism (RFLP). The total nosZ RFLP profiles increased in similarity with soil sample size until triplicate 3-g samples produced visually identical RFLP profiles for each treatment. Large differences in total nosZ profiles were observed between the native and cultivated soils. The fragments representing major groups of clones encountered at least twice and four randomly selected clones with unique RFLP patterns were sequenced to verify nosZ identity. The sequence diversity of nosZ clones from the cultivated field was higher, and only eight patterns were found in clone libraries from both soils among the 182 distinct nosZ RFLP patterns identified from the two soils. A group of clones that comprised 32% of all clones dominated the gene library of native soil, whereas many minor groups were observed in the gene library of cultivated soil. The 95% confidence intervals of the Chao1 nonparametric richness estimator for nosZ RFLP data did not overlap, indicating that the levels of species richness are significantly different in the two soils, the cultivated soil having higher diversity. Phylogenetic analysis of deduced amino acid sequences grouped the majority of nosZ clones into an interleaved Michigan soil cluster whose cultured members are α-Proteobacteria. Only four nosZ sequences from cultivated soil and one from the native soil were related to sequences found in γ-Proteobacteria. Sequences from the native field formed a distinct, closely related cluster (D_mean = 0.16) containing 91.6% of the native clones. Clones from the cultivated field were more distantly related to each other (D_mean = 0.26), and 65% were found outside of the cluster from the native soil, further indicating a difference in the two communities. Overall, there appears to be a relationship between use and richness, diversity, and the phylogenetic position of nosZ sequences, indicating that agricultural use of soil caused a shift to a more diverse denitrifying community.     

Influence of polymeric 3-alkylpyridinium salts from the marine sponge Reniera sarai on the growth of algae and wood decay fungi

Polymeric alkylpyridinium salts (poly-APS) isolated from the marine sponge Reniera sarai act as antifouling and anticholinesterase agents. They also show moderate haemolytic and cytotoxic activities against different cell lines. The haemolytic activity of poly-APS is due to their detergent-like structure and behaviour in aqueous solutions. In this work, the lytic activity of poly-APS against freshwater and marine algae, and inhibitory effects on wood decay fungi, were investigated. The results show that poly-APS inhibit the proliferation and movements of susceptible algae. Effects of poly-APS were time- and concentration-dependent and differed between various algal species. No growth inhibition effects were observed towards the examined wood fungi.     

INTERCOLONIAL VARIABILITY IN MACROMOLECULAR COMPOSITION IN P‐STARVED AND P‐REPLETE SCENEDESMUS POPULATIONS REVEALED BY INFRARED MICROSPECTROSCOPY1

Macromolecular variability in microalgal populations subject to different nutrient environments was investigated, using the chlorophyte alga Scenedesmus quadricauda (Turpin) Bréb. as a model organism. The large size of the four‐cell coenobia in the strain used in this study (～35 μm diameter) conveniently allowed high quality spectra to be obtained from individual coenobia using a laboratory‐based Fourier transform infrared (FTIR) microscope with a conventional globar source of IR. By drawing sizable subpopulations of coenobia from two Scenedesmus cultures grown under either nutrient‐replete or P‐starved conditions, the population variability in macromolecular composition, and the effects of nutrient change upon this, could be estimated. On average, P‐starved coenobia had higher carbohydrate and lower protein absorbance compared with P‐replete coenobia. These parameters varied between coenobia with histograms of the ratio of absorbance of the largest protein and carbohydrate bands being Gaussian distributed. Distributions for the P‐replete and P‐starved subpopulations were nonoverlapping, with the difference in mean ratios for the two populations being statistically significant. Greater variance was observed in the P‐starved subpopulation. In addition, multivariate models were developed using the spectral data, which could accurately predict the nutrient status of an independent individual coenobium, based on its FTIR spectrum. Partial least squares discriminant analysis (PLS‐DA) was a better prediction method compared with soft independent modeling by class analogy (SIMCA).     

Some minerals as well as trace and toxic elements in livers of fallow deer (<Emphasis Type="Italic">Dama dama</Emphasis>) in Slovenia

Minerals were determined in the livers of 69 fallow deer, which were shot during routine culls in Slovenian enclosures, in the winter of 2001/2002. Analysis of minerals and trace elements was done by atomic absorption spectroscopy and analysis of phosphorus was done by spectrophotometry. Mean values recorded for liver were 0.11±0.07 g/kg for calcium, 6.38±1.10 g/kg for phosphorus, 0.65±0.24 g/kg for sodium, 1.91±0.66 g/kg for potassium, 0.14±0.03 g/kg for magnesium, 31.0±21.9 mg/kg for copper, 31.3±10.8 mg/kg for zinc, 3.8±1.7 mg/kg for manganese, and 141±65 mg/kg for iron. Lead was present in high concentrations in the liver of almost all animals (1.48±0.92 mg/kg), while cadmium occurred in relatively low concentrations (0.27±0.14 mg/kg). There were significant differences in liver sodium, copper and iron concentrations between sexes and significant differences in liver phosphorus concentration between old and young animals.     

Inverted sinonasal papilloma--a report of 31 cases and review of the literature

Inverted papilloma is an epithelial neoplasm of the lateral nasal wall and adjacent sinuses characterized by a marked propensity for recurrence and a significant association with carcinoma. In this retrospective study we present 31 cases treated by our departments between 1982 and 1999. The aim was to compare our results to those of other authors especially regarding surgical management. The male to female ratio of these patients was 2:1 and most patients were in the 6th and 7th decades of life. Conservative surgery was used in most cases as the initial treatment. The overall recurrence rate was low and there were 3 cases associated with carcinoma. We conclude that the results of conservative surgery in selected cases are comparable to those using radical methods. A review of the literature is presented and particular attention is dedicated to the literature concerning analysis of p53 expression, HPV and Epstein-Barr infection and apoptosis in inverted papilloma.     

The effect of activation of Mammalian oocytes on remodeling of donor nuclei after nuclear transfer

Activation of bovine oocytes by experimental procedures that closely mimic normal fertilization is essential both for intracytoplasmic sperm injection and for nuclear transfer (NT). Therefore, with the goal of producing haploid activated oocytes, we evaluated whether butyrolactone I and bohemine, either alone or in combination with ionomycin, are able to activate young matured mammalian oocytes. Furthermore, the effect on the patterns of DNA synthesis after pronuclear formation as well as changes in histone H1 kinase and MAP kinase activities during the process of activation were studied. Our results with bohemine show that the specific inhibition of cyclin-dependent kinases (CDKs) in metaphase II bovine oocytes induces parthenogenetic activation in a dose dependent manner (25, 50, and 100 microM, respectively), either alone (3%, 30%, and 50%) or in combination with ionomycin (30%, 70%, and 87.5%). The effect of two activation protocols on nuclear remodeling, DNA synthesis during the first cell cycle, chromosome segregation after first mitosis, and development to blastocyst of embryos produced by somatic nuclear transfer were studied. Pronuclear formation was significantly higher when activation lasted 5 h compared to 3 h for both ethanol-cycloheximide and ionomycin-bohemine treatment. Initiation of DNA synthesis was delayed in ethanol-cycloheximide group, however, after 12-h labeling 100% of embryos synthesized DNA in both groups. Analysis of two-cell embryos with DNA probes for chromosome 6, 7, and 15 by fluorescence in situ hybridization showed that at least 50% of NT embryos were of normal ploidy, independent of the activation protocol.     

Endoscopic Treatment of Biliary Stenosis in Patients with Alveolar Echinococcosis – Report of 7 Consecutive Patients with Serial ERC Approach

Background and Aims

Biliary vessel pathology due to alveolar echicococcosis (AE) results in variable combinations of stenosis, necrosis and inflammation. Modern management strategies for patients with cholestasis are desperately needed. The aim is proof of principle of serial ERC (endoscopic retrograde cholangiography) balloon dilation for AE biliary pathology.

Methods

Retrospective case series of seven consecutive patients with AE-associated biliary pathology and ERC treatment in an interdisciplinary endoscopy unit at a University Hospital which hosts a national echinococcosis treatment center. The AE patient cohort consists of 106 patients with AE of the liver of which 13 presented with cholestasis. 6/13 received bilio-digestive anastomosis and 7/13 patients were treated by ERC and are reported here. Biliary stricture balloon dilation was performed with 18-Fr balloons at the initial and with 24-Fr balloons at subsequent interventions. If indicated 10 Fr plastic stents were placed.

Results

Six patients were treated by repeated balloon dilation and stenting, one by stenting only. After an acute phase of 6 months with repeated balloon dilation, three patients showed “sustained clinical success” and four patients “assisted therapeutic success,” of which one has not yet reached the six month endpoint. In one patient, sustained success could not be achieved despite repeated insertion of plastic stents and balloon dilation, but with temporary insertion of a fully covered self-expanding metal stent (FCSEMS). There was no loss to follow up. No major complications were observed.

Conclusions

Serial endoscopic dilation is a standard tool in the treatment of benign biliary strictures. Serial endoscopic intervention with balloon dilation combined with benzimidazole treatment can re-establish and maintain biliary duct patency in AE associated pathology and probably contributes to avoid or postpone bilio-digestive anastomosis. This approach is in accordance with current ERC guidelines and is minimally disruptive for patients.     

The role of microcystins in heavy cyanobacterial bloom formation

The presence of high microcystin concentrations in cyanobacterialblooms additionally affects species diversity. Blooms with hightoxin contents can reach higher cell densities, which is alsodemonstrated by microcystin cell contents. In vitro experimentsshow that microcystins influence phytoplankton proliferation.The action is strongly dependent on the phytoplankton speciestested and light conditions. We propose that the environmentalimpact of different microcystins depends on their enzymaticinhibition activity and thus could not be measured merely onthe basis of their toxicity to vertebrate species. Their rolein heavy cyanobacterial bloom and scum formation is discussed,as well as their impact on the massive proliferation of otherspecies following toxic cyanobacterial bloom degradation.     

Comparison of ultrasound-guided vs laparoscopic transvaginal ovum pick-up (OPU) in simmental heifers

Transvaginal ovum pick-up (OPU) offers several advantages over standard embryo transfer procedures. For a systematic comparison of the ultrasound-guided (U-OPU) and the laparoscopic OPU (L-OPU) method, groups of Simmental heifers were subjected to 1 of the 2 OPU-methods for 8 wk (15 sessions) followed by a treatment-free interval of 11 wk and then another 8-wk period of OPU using the alternative method. Parameters that were evaluated included the number of follicles aspirated, number and morphology of recovered cumulus-oocyte complexes (COCs), and developmental capacity of oocytes after in vitro maturation (IVM) and fertilization (IVF). Blood samples were also taken from the donors twice a wk for progesterone measurement. To evaluate effects of long-term OPU on subsequent fertility of donors, all heifers (n = 14) were inseminated during the first natural estrus after the OPU experiment. The proportion of Class I oocytes was significantly (P < 0.001) higher after U-OPU than after L-OPU (38.7% vs 21.0%). Following IVM/IVF, this difference in oocyte quality was reflected by the cleavage rate (U-OPU: 58.1%; L-OPU: 52.1%; P < 0.05) and the rate of development to morulae and blastocysts (U-OPU: 27.1%; L-OPU: 13.9%; P < 0.001). Among other factors, the greater changes in vacuum pressure during L-OPU vs U-OPU might be responsible for the difference in oocyte quality. This problem may be overcome by a more flexible system for regulating the vacuum. Progesterone levels were higher during the L-OPU than the U-OPU periods. Seven donors (50%) were diagnosed pregnant by ultrasonography on Day 28 and by palpation per rectum on Day 42.