Colonization of termite hindgut walls by oxymonad flagellates and prokaryotes in Incisitermes tabogae,I. marginipennis and Reticulitermes flavipes

We studied the colonization of the paunch wall of three lower termites, Reticulitermes flavipes, Incisitermes tabogae, and Incisitermes marginipennis, by light and electron microscopy. In addition to various prokaryotes, oxymonad flagellates were attached to the wall of the paunch in all three species. The prokaryotic layer found in R. flavipes is relatively thin, since most organisms are attached laterally. Large members of the flagellate genus Pyrsonympha protrude into the gut lumen. The prokaryotes are very abundant on the gut wall in I. tabogae and I. marginipennis, forming a thick carpet of mostly vertically attached rods and wavy spirochetes. The adhering oxymonads are relatively small and almost hidden in the thick bacterial biofilm. Three small morphotypes were seen in I. tabogae; two possessing a short rostellum and one amoeboid. The only oxymonad found in I. tabogae so far, Oxymonas clevelandi, is not identical to any of the present oxymonads. I. marginipennis contains a mid-sized oxymonad with ectobiotic spirochetes, probably identical to Oxymonas hubbardi, and a tiny unknown morphotype. The spatial organization of the pro- and eukaryotic microorganisms on the gut wall of the three termites is described and discussed concerning oxygen stress.     

Comparison of five different scoring methods in the evaluation of inflammatory infiltration (tumor‐infiltrating lymphocytes) in superficial spreading and nodular melanoma

The objective of our study was to compare the five different scoring methods of tumor‐infiltrating lymphocytes (TILs) assessment in a group of 213 cases of superficial spreading and nodular melanoma. The scoring methods include (a) Clark scoring; (b) Melanoma Institute Australia system; (c) scoring system used in the study of Saldanha et al.; (d) scoring system used in the TCGA study and modified by Park et al.; and (e) the system recently proposed by the “International Immuno‐Oncology Biomarker Working Group” for TILs scoring in all solid tumors. Prediction of survival with three main outcomes—disease‐specific‐free survival, local recurrence‐free survival, and distant metastasis‐free survival—was evaluated. The prognostic value of TILs showed statistical significance in univariate analysis regarding all three of the outcomes only for three of the five evaluated methods; the Clark scoring, the Melanoma Institute Australia system, and the system proposed by the “International Immuno‐Oncology Biomarker Working Group”. However, in multivariate analysis with covariants including Breslow thickness, type of melanoma, location, sex, and age, we did not find TILs to be an independent prognostic factor.     

Effects of Bark Beetle Disturbance on Soil Nutrient Retention and Lake Chemistry in Glacial Catchment

Ecosystems - Forest ecosystems worldwide are subjected to human-induced stressors, including eutrophication and acidification, and to natural disturbances (for example, insect infestation,...     

Red-shifted light-harvesting system of freshwater eukaryotic alga Trachydiscus minutus (Eustigmatophyta,Stramenopila)

Photosynthesis Research - Survival of phototrophic organisms depends on their ability to collect and convert enough light energy to support their metabolism. Phototrophs can extend their absorption...     

Time course of cortical and zona reactions of pig oocytes upon intracellular calcium increase induced by thimerosal

Our previous study indicated that thimerosal is one of the most effective artificial activators to mimic sperm-induced increases in the intracellular free calcium concentration ([Ca2+]i) and other activation events in pig oocytes (Macháty et al., 1997). The present study was conducted to examine the temporal relationship between intracellular calcium transients, cortical granule (CG) exocytosis and the zona reaction induced by thimerosal. When pig oocytes matured in vitro were exposed to 200 microM thimerosal the first intracellular calcium transient, with a mean peak ratio of 4.97 +/- 1.14, was observed 509.64 +/- 122.03 s after addition of thimerosal. The density of CGs fell significantly from 63.3 +/- 11.7 CGs/100 micron 2 of cortex in control oocytes to 25.7 +/- 19.2 CGs/100 micron 2 of cortex (59.4% release) at 2 min after the first intracellular calcium transient. At 5 min after the calcium transient the residual CG density had been reduced to 10.7 +/- 10.4 CGs/100 micron 2 of cortex (83.1% release). This degree of CG exocytosis was the same as that in oocytes penetrated by sperm (9.5 +/- 5.1 CGs/100 micron 2 of cortex). No further decrease in residual CG density was observed at 10 min (10.3 +/- 14.8 CGs/100 micron 2 of cortex). Whereas 77.4% (120/155) of control oocytes were penetrated by spermatozoa only 1.4% (2/144) of thimerosal-treated oocytes were penetrated. Further experimental results obtained by in vitro fertilisation of oocytes with preincubated (capacitated) spermatozoa suggested that the zona block to sperm penetration in thimerosal-treated oocytes occurred within 35 min after CG exocytosis and 40 min after the first calcium transient. These results indicate that polyspermic penetration of pig oocytes inseminated in vitro is not due to delayed or incomplete CG exocytosis but more likely to a delayed zona reaction and/or simultaneous sperm penetration.     

Development of pig embryos by nuclear transfer of cultured fibroblast cells

Pig fibroblast cells were transferred to enucleated oocytes by micromanipulation and electrofusion. The donor cells used for nuclear transfer were synchronized in presumptive G0 by serum starvation. In the first experiment, nuclear transfer was performed with fibroblasts that had either a smooth or a rough surface. A significant difference (p < 0.05) in the percentage of chromosome condensation (39.5%, 15/38 and 16.6%, 5/30) and nuclear formation (36.8%, 14/38 and 16.3%, 8/49) was found between the reconstructed embryos derived from the cells with smooth surface and with rough surfaces, respectively. The percentage of chromosome condensation (42.5%, 17/40 and 19.6%, 11/56) and nuclear formation (38.3%, 23/60 and 18.8%, 9/48) were higher (p < 0.05) in reconstructed embryos derived from small (15 microm) donor cells compared to large donor cells (20 microm), respectively. The percentage nuclei at 3 different time points (3, 6, and 9 hours in culture medium) was higher (p = 0.003) in the reconstructed embryos activated by thimerosal and dithiothreitol (20%, 36%, and 41.3%) compared to those without activation treatment (0%, 11.8%, and 22.2%). In addition, there was an increased percentage with nuclei as the time in culture increased from 3 to 9 hours (p = 0.029). The percentages of chromosome condensation (34.6%; 9/26) and nuclear formation (33.3%; 9/27) in nuclear transfer embryos were similar. The rate of blastocysts/morulae development (14.0%; 6/43) was low. However, 2 cavitated embryos (presumptive blastocysts) with 14 and 11 nuclei and 1 morula with 8 nuclei were obtained. This together with the above evidence indicate that the nuclei from pig fibroblast cells can be partially reprogrammed, which suggests that transfer of nuclei from fibroblast cells to in vitro matured oocytes resulting in production of identical or genetically altered pigs may be possible.     

Parthenogenetic activation of porcine oocytes after nuclear transfer

Mature porcine oocytes are arrested at metaphase II of meiosis. At fertilization, like all mammalian oocytes they exhibit a low frequency Ca(2+) oscillation lasting several hours. This oscillation is thought to be the signal that triggers resumption of meiosis and activates the developmental program of the oocyte. The signal transduction mechanism of the sperm-induced Ca(2+) signal is not known in detail, and attempts to generate the oscillation artificially have met with little success. Nevertheless, artificial activation of the oocyte is a crucial step during nuclear transfer. Methods are available to induce a transient elevation in the intracellular free Ca(2+) concentration to surpass the meiotic arrest and induce development of the constructed embryo. Further studies concentrating on the mechanism of Ca(2+) signaling during fertilization will help to improve the efficiency of the procedures used for parthenogenetic activation of the oocyte.