Genetic lesions within the 3a gene of SARS-CoV

Phage N5 is one of the phages of Vibrio cholerae serovar O1 biotype El Tor (Ghosh, A. N., Ansari, M. Q., and Dutta, G. C. Isolation and morphological characterization of El Tor cholera phages. J. Gen. Virol. 70: 2241–2243, 1989). In the present communication the growth curve, molecular weight and confirmation of the genome, partial denaturation map and restriction endonuclease digestion pattern have been determined. Partial denaturation map indicates that the genome has non-permuted / invariant sequence. Presence of cohesive ends has also been documented.     

Failure to Use Cubicles and Concentrate Dispenser by Heifers after Transfer from Rearing Accommodation to Milking Herd

Thirty-three dairy farms in the Norwegian counties of ?stfold and Akershus in which cubicle sheds had been in use for at least one year and with a herd size of less than 60 cows, were contacted and asked to participate in a study. The study focused on heifers' use of cubicles and concentrate dispenser just after being transferred from rearing accommodation to the milking herd. For each heifer, the farmer recorded cubicle use once nightly between 9 and 11 pm. The daily amount of concentrate released in the dispenser and the allotted daily ration were also recorded. The recording period was 15 consecutive days for cubicle use and 7 days for concentrate dispenser use. Cubicle refusal behaviour, i.e. lying outside the cubicles, was analysed by logistic regression using rearing accommodation of heifers, herd size, heifer age, and housing layout as independent variables, and herd as a clustering variable. On Day 2 after transfer, 34% of the heifers were showing cubicle refusal behaviour (N = 340). By Day 15 this percentage had dropped to 23. Cubicle refusal was lower throughout the whole period among heifers which used the cubicles on the 3 first days after transfer compared to those which did not. This tendency could also be detected several months later. The analysis showed cubicle refusal to be significantly associated with rearing accommodation (OR = 6.1, c.i._95%OR = 1.5–24.3, P = 0.01) and cubicle layout in the shed (OR = 0.2, c.i._95%OR = 0.0–0.7, P = 0.01). None of the tested variables were found to be significant for failure to use the concentrate dispenser, a behaviour which was less frequent than cubicle refusal. However, 8 percent of the heifers did not visit the dispenser at all throughout the 7 days of observation.     

Impact of KCNE subunits on KCNQ1 (Kv7.1) channel membrane surface targeting

The KCNQ1 (Kv7.1) channel plays an important role in cardiovascular physiology. Cardiomyocytes co‐express KCNQ1 with KCNE1‐5 proteins. KCNQ1 may co‐associate with multiple KCNE regulatory subunits to generate different biophysically and pharmacologically distinct channels. Increasing evidence indicates that the location and targeting of channels are important determinants of their function. In this context, the presence of K⁺ channels in sphingolipid–cholesterol‐enriched membrane microdomains (lipid rafts) is under investigation. Lipid rafts are important for cardiovascular functioning. We aimed to determine whether KCNE subunits modify the localization and targeting of KCNQ1 channels in lipid rafts microdomains. HEK‐293 cells were transiently transfected with KCNQ1 and KCNE1–5, and their traffic and presence in lipid rafts were analyzed. Only KCNQ1 and KCNE3, when expressed alone, co‐localized in raft fractions. In addition, while KCNE2 and KCNE5 notably stained the cell surface, KCNQ1 and the rest of the KCNEs showed strong intracellular retention. KCNQ1 targets multiple membrane surface microdomains upon association with KCNE peptides. Thus, while KCNQ1/KCNE1 and KCNQ1/KCNE2 channels target lipid rafts, KCNQ1 associated with KCNE3–5 did not. Channel membrane dynamics, analyzed by fluorescence recovery after photobleaching (FRAP) experiments, further supported these results. In conclusion, the trafficking and targeting pattern of KCNQ1 can be influenced by its association with KCNEs. Since KCNQ1 is crucial for cardiovascular physiology, the temporal and spatial regulations that different KCNE subunits may confer to the channels could have a dramatic impact on membrane electrical activity and putative endocrine regulation. J. Cell. Physiol. 225: 692–700, 2010. © 2010 Wiley‐Liss, Inc.     

Morphological and molecular evidence for hybridization and introgression in Central European Arctium (Asteraceae)

Three species of burdock (Arctium lappa, A. tomentosum, and A. minus; Asteraceae), which commonly occur in sympatry in Central Europe, are thought to hybridize frequently, hence raising questions about their integrity. We surveyed mixed, monotypic, and/or single plant stands of these species in Central Germany to assess their extent of hybridization and introgression, and also the likelihood of insect host shifts between them. Individual plants were characterized by multivariate analyses of morphological and RAPD variation in combination with RAPD-based character-index-scores. All analyses recognized the three species as distinct units, but also identified a limited number of first (or early) generation hybrids between sympatric A. lappa and either A. tomentosum or A. minus, respectively, as well as one introgressant of A. tomentosum into A. lappa. While pre-zygotic isolation may partly account for the rarity of first generation crosses (F₁'s), the near absence of later generation hybrid segregants or introgressants likely reflects post-zygotic isolation due to the disruption of co-adapted gene complexes and/or habitat-mediated superiority shared by the ruderal parentals. As the tephritid fly Tephritis bardanae oviposits on both A. tomentosum (T) and A. minus (M), but not on A. lappa (L), the occurrence of L × M and L × T hybrids and/or introgressants may have important evolutionary consequences regarding the acquisition of a new host, i.e. A. lappa, by these insects.     

Chloroplast DNA and isozyme analysis of the progenitor-derivative species relationship between Senecio nebrodensis and S. viscosus (Asteraceae)

A comparison of chloroplast DNA (cpDNA) and isozyme diversity in Senecio nebrodensis L., a species restricted to certain mountain ranges in Spain, and S. viscosus L., a widespread ruderal species in Europe, revealed that S. viscosus possessed the more common of two cpDNA haplotypes resolved in S. nebrodensis and contained only a small subset of the alleles found in S. nebrodensis at enzyme coding loci. The two species shared one restriction length mutation and one site mutation in their cpDNA, which distinguished them from other European Senecio species examined previously. Taken overall, these results support the hypothesis that S. nebrodensis and S. viscosus are related as a progenitor-derivative pair of species. The fact that no novel alleles were found in S. viscosus would suggest a relatively recent origin of the species, most probably in late glacial or postglacial times.     

Foetal liver infusion in a chronic myeloid leukemia patient with busulphan toxicity

A 36-year-old man suffering from chronic myeloid leukemia (in chronic phase) was initially treated with busulphan. At the end of 6 months of follow-up he developed bone marrow aplasia. He was given single foetal liver infusion therapy. The patient recovered fully from aplasia. He continued in chronic phase for more than 7 years with intermittent busulphan therapy.     

A comparative study on genetic effects of artificial and natural habitat fragmentation on Loropetalum chinense (Hamamelidaceae) in Southeast China

Elucidating the demographic and landscape features that determine the genetic effects of habitat fragmentation has become fundamental to research in conservation and evolutionary biology. Land-bridge islands provide ideal study areas for investigating the genetic effects of habitat fragmentation at different temporal and spatial scales. In this context, we compared patterns of nuclear microsatellite variation between insular populations of a shrub of evergreen broad-leaved forest, Loropetalum chinense, from the artificially created Thousand-Island Lake (TIL) and the Holocene-dated Zhoushan Archipelago of Southeast China. Populations from the TIL region harboured higher levels of genetic diversity than those from the Zhoushan Archipelago, but these differences were not significant. There was no correlation between genetic diversity and most island features, excepting a negative effect of mainland–island distance on allelic richness and expected heterozygosity in the Zhoushan Archipelago. In general, levels of gene flow among island populations were moderate to high, and tests of alternative models of population history strongly favoured a gene flow-drift model over a pure drift model in each region. In sum, our results showed no obvious genetic effects of habitat fragmentation due to recent (artificial) or past (natural) island formation. Rather, they highlight the importance of gene flow (most likely via seed) in maintaining genetic variation and preventing inter-population differentiation in the face of habitat ‘insularization'' at different temporal and spatial scales.     

Apoptosis mapping in space and time of 3D tumor ecosystems reveals transmissibility of cytotoxic cancer death

The emerging tumor-on-chip (ToC) approaches allow to address biomedical questions out of reach with classical cell culture techniques: in biomimetic 3D hydrogels they partially reconstitute ex vivo the complexity of the tumor microenvironment and the cellular dynamics involving multiple cell types (cancer cells, immune cells, fibroblasts, etc.). However, a clear bottleneck is the extraction and interpretation of the rich biological information contained, sometime hidden, in the cell co-culture videos. In this work, we develop and apply novel video analysis algorithms to automatically measure the cytotoxic effects on human cancer cells (lung and breast) induced either by doxorubicin chemotherapy drug or by autologous tumor-infiltrating cytotoxic T lymphocytes (CTL). A live fluorescent dye (red) is used to selectively pre-stain the cancer cells before co-cultures and a live fluorescent reporter for caspase activity (green) is used to monitor apoptotic cell death. The here described open-source computational method, named STAMP (spatiotemporal apoptosis mapper), extracts the temporal kinetics and the spatial maps of cancer death, by localizing and tracking cancer cells in the red channel, and by counting the red to green transition signals, over 2–3 days. The robustness and versatility of the method is demonstrated by its application to different cell models and co-culture combinations. Noteworthy, this approach reveals the strong contribution of primary cancer-associated fibroblasts (CAFs) to breast cancer chemo-resistance, proving to be a powerful strategy to investigate intercellular cross-talks and drug resistance mechanisms. Moreover, we defined a new parameter, the ‘potential of death induction’, which is computed in time and in space to quantify the impact of dying cells on neighbor cells. We found that, contrary to natural death, cancer death induced by chemotherapy or by CTL is transmissible, in that it promotes the death of nearby cancer cells, suggesting the release of diffusible factors which amplify the initial cytotoxic stimulus.