Brilliant cresyl blue staining allows the selection for developmentally competent immature feline oocytes.

The outcome of in-vitro-maturation and in-vitro-fertilization of feline oocytes depends on the selection of high quality oocytes, and is often restricted to morphological criteria. The aim of this study was to test whether the Brilliant cresyl blue (BCB) staining is suitable for pre-selection of feline oocytes before in-vitro-maturation. Cumulus-oocytes-complexes (COC) were released from domestic cat ovaries obtained after ovariectomy and were subjected to BCB staining. BCB+ stained oocytes were characterized by a violet/pale blue staining of the ooplasma, BCB- oocytes remained unstained. Transmission electron microscopy indicated for a slightly advanced stage of BCB- oocytes within the maturation process. After 24 h in-vitro-maturation, almost 75% of BCB+ and 21.5% of BCB- oocytes were able to reach metaphase II. Also, after in-vitro-fertilization, significantly more oocytes developed to morulae (19.2%) if oocytes were preselected for BCB staining, although 8% of unstained COC still reached advanced embryo stages. Prolonged storage of ovaries before COC retrieval for 16-20 h at 4 °C was accompanied by reduced number of BCB+ oocytes (96 of 210, 45.7%) in comparison to freshly isolated COC (151 of 225, 67.1%), and impaired cleavage rate (19.8%) and morula rate (9.4%) of BCB+ oocytes but the rate of embryos which developed to advanced stages remained unchanged (∼50%). To conclude, BCB staining is a very useful tool to preselect immature COC of feline species ensuring higher developmental rates.

Assessing the toxicity of contaminated soils using the nematode Caenorhabditis elegans as test organism.

In this study, nine uncontaminated reference soils and 22 contaminated soils with different physico-chemical properties and contamination patterns were tested with a standardized toxicity test, using the nematode, Caenorhabditis elegans, as test organism. Fertility, growth and reproduction of C. elegans in the soils were compared with the exposure in standard soil Lufa St.2.2. C. elegans showed 100% fertility and a very low variability of growth in the reference soils. Although, reproduction varied considerably between the various reference soils, validity criteria (>30 offspring per test organism) were met in all reference soils. Moreover, Lufa St. 2.2 turned out to be a suitable and representative control soil. In order to clearly classify the effects of the polluted soils on C. elegans, toxicity thresholds were derived for nematode fertility (20% inhibition), growth (10% inhibition) and reproduction (40% inhibition) on the basis of the test inherent variability (MDD=minimal detectable difference), as well as their variability between the uncontaminated reference soils (MTI=maximal tolerable inhibition). The contaminated soils showed clear toxic effects on the nematodes, whereas the toxicity was better correlated to organic than to heavy metal contamination in bulk soil. Interestingly, the results of the nematode toxicity test were not well correlated with those of tests with oligochaetes, collembolans and plants, performed with the same soils, showing that the results are not redundant. The toxicity test using C. elegans turned out to be suitable for testing the toxicity of field collected soils and might by a valuable addition to soil test batteries.

The effect of tributyltin-oxide on earthworms, springtails, and plants in artificial and natural soils.

Chemical bioavailability in Organisation for Economic Co-operation and Development (OECD) artificial soil can contrast with bioavailability in natural soils and produce ecotoxicologic benchmarks that are not representative of species' exposure conditions in the field. Initially, reproduction and growth of earthworm and Collembolan species, and early seedling growth of a dicotyledonous plant species, in nine natural soils (with a wide range of physicochemical properties) and in OECD soil were evaluated. Soils that supported reproduction and growth of the test species were then used to investigate the toxicity of tributyltin-oxide (TBT-O). Natural soils caused greater toxicity of TBT-O to earthworms (EC(50) values varied from 0.5 to 4.7 mg/kg soil dry weight [dw]) compared with toxicity in OECD soil (EC(50) = 13.4 mg/kg dw). Collembolans were less sensitive to TBT-O than earthworms in natural soils, with EC(50) values ranging from 23.4 to 177.8 mg/kg dw. In contrast, the toxicity of TBT-O to collembolans in OECD soil (EC(50) = 104.0 mg/kg dw) was within the range of EC(50) values in natural soils. Phytotoxicity tests revealed even greater difference between the effects in natural soils (EC(50) values ranged from 10.7 to 189.2 mg/kg dw) and in OECD soil (EC(50) = 535.5 mg/kg dw) compared with results of the earthworm tests. Studies also showed that EC(50) values were a more robust end point compared with EC(10) values based on comparisons of coefficients of variation. These results show that toxicity testing should include studies with natural soils in addition to OECD soil to better reflect exposure conditions in the field.

The use of earthworms in ecological soil classification and assessment concepts.

Without doubt, earthworms are the most important soil invertebrates in most soils worldwide, in terms of both biomass and activity. Several species are even considered to be ecosystem engineers. Earthworms are also known to influence soil structure, soil chemistry, and, in particular, processes like organic matter decomposition. In addition, standardized sampling methods are available and their taxonomy is well known (even the first PC-aided keys have been developed). For these reasons, earthworms were recognized as a part of ecological classification and assessment schemes early on. However, due to the relatively small number at many sites, they have to be part of a battery approach. By use of examples from The Netherlands (biological indicator of soil quality) and Germany (soil biological site classification), the practicability of the use of earthworms is demonstrated in determining the influence of different anthropogenic land use forms. In these cases, the structure of the earthworm community, as well as their abundance and biomass, were used as endpoints.

The use of enchytraeids in ecological soil classification and assessment concepts.

In many soils worldwide, enchytraeids (Annelida: Oligochaeta) play a major role in soil functions like the decomposition of organic matter. In addition, standardized sampling methods are available and taxonomic as well as ecological knowledge is rapidly increasing (in particular, ecological profiles of the most abundant species). For these reasons, enchytraeids were recognized as a part of ecological classification and assessment schemes. In this contribution, the most important of these schemes are presented and it is shown that enchytraeids are most valuable as part of a battery approach. By use of examples from The Netherlands (BISQ), and Germany (BBSK), the practicability of the use of enchytraeids is demonstrated in identifying the influence of different anthropogenic land use forms. In these cases, the structure of the enchytraeid community and their abundance and biomass were used as endpoints. However, automation of the identification process can still be improved.

Structural chemistry of polycyclic heteroaromatic compounds. Part XI. Photoelectron spectra and electronic structures of tetracyclic hetarenes of the triphenylene type.

The UV photoelectron spectra of several tetracyclic heteroaromatic compounds (2-9) which are pi-isoelectronic with triphenylene (1) have been recorded and analysed making use of semiempirical AM1 and PM3 as well as ab initio/DFT B3LYP calculations. In one series of compounds (2-7), the peripheral benzene rings of 1 are successively substituted by thiophene rings that are either [b]- or [c]-annellated with the central benzene unit. In 2-7 only marginal shifts are found for most of the IPs of electrons. In the benzotrithiophenes 5-7, a systematic variation is displayed by IP(pi7). Compared to 1, the pi electron system of benzo[c]trithiophene (7) is approximately two times as much destabilized as in the isomers 5 and 6 with [b]annellated thiophene rings. The IP[n(S)] values of the thiophene derivatives 2-7 indicate that these orbitals are clearly destabilized relative to thiophene. The same holds for the n(O) orbital of the furane derivative 9 in comparison with that of furane. In 9, only the higher pi MOs (pi7-pi9) are destabilized whereas the lower levels (pi1-pi4) are stabilized, and those in between (pi5-pi6) remain essentially unshifted. In the pyrrole derivative 8, all pi MOs are substantially destabilized by about 0.5-1.6 eV relative to 1.