Developmental competence of bovine oocytes: effects of follicle size and the phase of follicular wave on in vitro embryo production.

Abstract Developmental competence of bovine oocytes collected from follicles of different size categories (in either the growth or the dominant phase of the first follicular wave) was studied, with the aim of improving in vitro embryo production. Estrus and ovulation of 39 cyclic Holstein dairy cows were synchronized by two prostaglandin F 2a treatments at 11-day intervals and one hCG treatment on the day of onset of estrus (Day 0). Cows with follicles in either the growth (Day 3, n ¼ 25) or the dominant phase (Day 7, n ¼ 14) were slaughtered, and follicles >5 mm were counted. Three oocyte populations were recovered separately from large (11-15 mm), medium (6-10 mm) and small (2-5 mm) follicles in both follicular phases. All collected cumulus-oocyte complexes (COC), except for markedly atretic oocytes without cumulus cells, were used in experiments. Oocytes were matured, fertilized and cultured by standard methods. There were no significant differences between the growth and the dominant phases for mean numbers of large follicles, useable oocytes and embryos per donor. Generally, those numbers were low, but the development rates of oocytes into blastocysts were high, particularly in the growth phase (60.0%). Mean (AES.E.M.) numbers of medium follicles, oocytes and embryos per donor were higher in the growth as compared with the dominant phase; in the useable oocytes and embryos, this difference was significant (9:6 AE 1:4 and 3:5 AE 0:6 versus 3:9 AE 0:6 and 1:1 AE 0:3; P < 0:01). The development rates of oocytes into blastocysts, however, did not differ significantly between the growth and the dominant phases (36.7% versus 27.8%). Mean numbers of useable oocytes and embryos per donor recovered from small follicles in both follicular wave phases were similar. The development rate of oocytes into blastocysts was generally low, but higher (P < 0:01) in the growth than in the dominant phase (24.5% versus 11.7%). Comparison between the two phases showed that mean number of all counted follicles and all useable oocytes collected per donor were similar, but the mean number of embryos per donor and the development rate of oocytes into blastocysts were higher in the growth phase than in the dominant phase (8:0 AE 1:2 versus 3:8 AE 2:4; P ¼ 0:012 and 30.3% versus 14.9%; P < 0:01). The interaction between follicle size and the phase of follicular wave affected the efficiency of embryo production. The yield of embryos was primarily influenced by the number of oocytes collected from medium follicles and the developmental competence of oocytes from small follicles. The growth phase was more effective for oocyte collection; the number of oocytes from medium follicles and the developmental competence of oocytes from small follicles decreased in the dominant phase.

The Effects of Mary Rose Conservation Treatment on Iron Oxidation Processes and Microbial Communities Contributing to Acid Production in Marine Archaeological Timbers

The Tudor warship the Mary Rose has reached an important transition point in her conservation. The 19 year long process of spraying with polyethylene glycol (PEG) has been completed (April 29th 2013) and the hull is air drying under tightly controlled conditions. Acidophilic bacteria capable of oxidising iron and sulfur have been previously identified and enriched from unpreserved timbers of the Mary Rose, demonstrating that biological pathways of iron and sulfur oxidization existed potentially in this wood, before preservation with PEG. This study was designed to establish if the recycled PEG spray system was a reservoir of microorganisms capable of iron and sulfur oxidization during preservation of the Mary Rose. Microbial enrichments derived from PEG impregnated biofilm collected from underneath the Mary Rose hull, were examined to better understand the processes of cycling of iron. X-ray absorption spectroscopy was utilised to demonstrate the biological contribution to production of sulfuric acid in the wood. Using molecular microbiological techniques to examine these enrichment cultures, PEG was found to mediate a shift in the microbial community from a co-culture of Stenotrophomonas and Brevunidimonas sp, to a co-culture of Stenotrophomonas and the iron oxidising Alicyclobacillus sp. Evidence is presented that PEG is not an inert substance in relation to the redox cycling of iron. This is the first demonstration that solutions of PEG used in the conservation of the Mary Rose are promoting the oxidation of ferrous iron in acidic solutions, in which spontaneous abiotic oxidation does not occur in water. Critically, these results suggest PEG mediated redox cycling of iron between valence states in solutions of 75% PEG 200 and 50% PEG 2000 (v/v) at pH 3.0, with serious implications for the future use of PEG as a conservation material of iron rich wooden archaeological artefacts

Lawsonia intracellularis exploits β-catenin/Wnt and Notch signalling pathways during infection of intestinal crypt to alter cell homeostasis and promote cell proliferation

Lawsonia intracellularis is an obligate intracellular bacterial pathogen that causes proliferative enteropathy (PE) in pigs. L. intracellularis infection causes extensive intestinal crypt cell proliferation and inhibits secretory and absorptive cell differentiation. However, the affected host upstream cellular pathways leading to PE are still unknown. β-catenin/Wnt signalling is essential in maintaining intestinal stem cell (ISC) proliferation and self-renewal capacity, while Notch signalling governs differentiation of secretory and absorptive lineage specification. Therefore, in this report we used immunofluorescence (IF) and quantitative reverse transcriptase PCR (RTqPCR) to examine β-catenin/Wnt and Notch-1 signalling levels in uninfected and L. intracellularis infected pig ileums at 3, 7, 14, 21 and 28 days post challenge (dpc). We found that while the significant increase in Ki67+ nuclei in crypts at the peak of L. intracellularis infection suggested enhanced cell proliferation, the expression of c-MYC and ASCL2, promoters of cell growth and ISC proliferation respectively, was down-regulated. Peak infection also coincided with enhanced cytosolic and membrane-associated β-catenin staining and induction of AXIN2 and SOX9 transcripts, both encoding negative regulators of β-catenin/Wnt signalling and suggesting a potential alteration to β-catenin/Wnt signalling levels, with differential regulation of the expression of its target genes. We found that induction of HES1 and OLFM4 and the down-regulation of ATOH1 transcript levels was consistent with the increased Notch-1 signalling in crypts at the peak of infection. Interestingly, the significant down-regulation of ATOH1 transcript levels coincided with the depletion of MUC2 expression at 14 dpc, consistent with the role of ATOH1 in promoting goblet cell maturation. The lack of significant change to LGR5 transcript levels at the peak of infection suggested that the crypt hyperplasia was not due to the expansion of ISC population. Overall, simultaneous induction of Notch-1 signalling and the attenuation of β-catenin/Wnt pathway appear to be associated with the inhibition of goblet cell maturation and enhanced crypt cell proliferation at the peak of L. intracellularis infection. Moreover, the apparent differential regulation of apoptosis between crypt and lumen cells together with the strong induction of Notch-1 signalling and the enhanced SOX9 expression along crypts 14 dpc suggest an expansion of actively dividing transit amplifying and/or absorptive progenitor cells and provide a potential basis for understanding the development and maintenance of PE

Charakterystyka popiołu dennego z MSWI oraz ocena odzysku

Municipal solid waste incineration (MSWI) bottom ash contains valuable components that can be recovered as secondary materials, such as ferrous and non-ferrous metals, some rare earth elements, glass etc. Metal-free mineral fraction can be used in construction industry as a substitute for natural materials. Important benefit of bottom ash recycling for the plant operator is also in reduction of fees for solid residuals landfilling. The composition of bottom ash is highly dependent on the composition of incinerated waste but in average can be around 5–13% ferrous metals, 2–5% non-ferrous metals, 15–30% glass and ceramics, 1–5% unburned organics and 50–70% mineral fraction. Several incineration plants in Europe are equipped with advanced systems for metals recovery, mostly based on magnetic separation of ferrous metals and separation of non-ferrous metals usually by eddy-current separators. To assess the possibilities of the bottom ash treatment in the Czech Republic it is necessary to obtain data about the bottom ash composition and evaluate its resource recovery potential. This paper summarizes characteristics of bottom ash samples from waste-to-energy plant in Prague. Emphasis of the study was primarily placed on the material composition. Bottom ash samples were dried and sieved into eight size fractions in the first step. It must be said that particle size distribution plays a decisive role for further utilization of bottom ash. In the second step, individual size fractions were sorted, using magnetic separation and the set of grinding, sieving, and manual separation processes, into the following materials: glass, ceramics and porcelain, magnetic particles with ferrous scrap, non-ferrous metals, unburned organic material, and residual fraction.Miejskie spalarnie odpadów stałych (ang. skrót MSWI) wytwarzają popiół, który zawiera cenne składniki, które można odzyskać w postaci materiałów wtórnych, tj. metali żelaznych i nieżelaznych, niektórych metali ziem rzadkich, szkła itd. Pozbawiona metalu frakcja mineralna może być użyta w przemyśle budowlanym jako zamiennik dla materiałów naturalnych. Ważną korzyścią płynącą z recyklingu popiołu dennego dla zarządzających spalarnią jest obniżenie kosztów składowania stałych pozostałości pospalaniu. Skład popiołu dennego w dużej mierze zależy od składu odpadów i średnio zawiera około 5-13% metali żelaznych, 2-5% metali nieżelaznych, 15-30% szkła i ceramiki, 1–5% niespalonych składników organicznych i 50-70% frakcji mineralnej. Kilka spalarni w Europie jest wyposażonych w zaawansowane systemy odzysku metali, głównie oparte o separacje magnetyczną. Aby ocenić możliwości odzysku popiołu dennego w Republice Czeskiej, zebrano dane na temat składu popiołu dennego i określono potencjał odzysku. Niniejsza praca podsumowuje charakterystykę próbek popiołu dennego pobranych ze spalarni generującej energię z odpadów znajdującej się w Pradze. Nacisk był przede wszystkim położony na skład materiału. W pierwszym etapie próbki popiołu dennego zostały osuszone i przesiane na 8 różnych frakcji. Warto uwzględnić, że rozkład wielkości ziaren ma decydujący wpływ na dalszą utylizację popiołu dennego. W drugim kroku, poszczególne frakcje zostały poddane separacji magnetycznej oraz innym procesom tj. rozdrabnianie, przesiewanie oraz separacja ręczna, na poszczególne frakcje: szkło, ceramika i porcelana, cząsteczki magnetyczne ze skrawkami żelaza, metale nieżelazne, niespalone materiały organiczne i pozostałe frakcje