Droplet-vitrification and morphohistological studies of cryopreserved shoot tips of cultivated and wild pineapple genotypes

Germplasm conservation of pineapple [Ananas comosus (L.) Mer.] is crucial to preserve the genus’ genetic diversity, to secure material for genetic improvement and to support innovative and new research. Long-term conservation is accomplished through cryopreservation, that is done by storing cells or tissues at ultra-low temperature in liquid nitrogen (−196 °C). Droplet-vitrification, a combination of droplet freezing and solution-based vitrification, was used to establish a protocol for cryopreservation of pineapple genetic resources. This protocol was tested on cultivated and wild pineapple genotypes to establish a long-term germplasm security duplicate as well as to investigate cryo-injuries in the tissues by means of histological techniques. Excised shoot tips (0.5–1 mm with one primordial leaf) of different pineapple genotypes were precultured for 48 h on solid MS medium containing 0.3 M of sucrose. Three PVS2 exposure times (30, 45 and 60 min) were tested. The results showed high post cryopreservation survival for all genotypes evaluated. The best PVS2 exposure time varied according to genotype, although 45 min gave the best survival for the majority of genotypes. The technique was highly efficient in cryopreserving meristem shoot tips of different pineapple genotypes, and was also less laborious than techniques previously reported. This is a first report on application of the droplet-vitrification technique to diverse genotypes of cultivated and wild pineapples and the first report on histological changes occurring in cryopreserved Ananas tissue.

     

Effects of prenatal exposure to a 900 MHz electromagnetic field on 60-day-old rat testis and epididymal sperm quality

We investigated the effects of exposure in utero to a 900 megahertz (MHz) electromagnetic field (EMF) on 60-day-old rat testis and epididymis. Pregnant rats were divided into control (CG; no treatment) and EMF (EMFG) groups. The EMFG was exposed to 900 MHz EMF for 1 h each day during days 13 ? 21 of pregnancy. Newborn rats were either newborn CG (NCG) or newborn EMF groups (NEMFG). On postnatal day 60, a testis and epididymis were removed from each animal. Epididymal semen quality, and lipid and DNA oxidation levels, apoptotic index and histopathological damage to the testis were compared. We found a higher apoptotic index, greater DNA oxidation levels and lower sperm motility and vitality in the NEMFG compared to controls. Immature germ cells in the seminiferous tubule lumen, and altered seminiferous tubule epithelium and seminiferous tubule structure also were observed in hematoxylin and eosin stained sections of NEMFG testis. Nuclear changes that indicated apoptosis were identified in TUNEL stained sections and large numbers of apoptotic cells were observed in most of the seminiferous tubule epithelium in the NEMFG. Sixty-day-old rat testes exposed to 900 MHz EMF exhibited altered sperm quality and biochemical characteristics.     

Rhodium Plasmonics for Deep-Ultraviolet Bio-Chemical Sensing

Rhodium (Rh) has been recently introduced as a perfect metal for ultraviolet (UV) applications with the advantages of its oxide-free nature and support of strong plasmon resonant modes at very short wavelengths. We report on a simple platform of nanoplasmonic structures to support strong plasmonic Fano resonances across the deep-UV spectrum for biochemical sensing applications. We investigate the plasmonic response of several types of Rh nanoparticles and designed dimer-type antennas using nanorings with geometrical tunability in both symmetric and antisymmetric assemblies. Using numerical and theoretical methods, it is shown that Rh-based dimer antennas with broken symmetry can be tailored to support strong plasmon resonant modes at the deep-UV region (\( E>6\; eV \)). We also propose a complex infinity-shaped structure composed of a pair of split rings with a nanodisk in between with extra degree of tunability to push the plasmon resonant modes further in deep-UV spectrum. Plasmon hybridization theory is used to describe formation of plasmonic Fano-resonant dips in simple nanoscale assemblies. We calculate the corresponding figure of merit for the Rh-based nanostructure around 11.5 which shows an excellent sensitivity to the refractive index perturbations of the surrounding medium at very short wavelengths for sensing applications.     

Molecular identification of 26 syntaxin genes and their assignment to the different trafficking pathways in Paramecium

SNARE proteins have been classified as vesicular (v)- and target (t)-SNAREs and play a central role in the various membrane interactions in eukaryotic cells. Based on the Paramecium genome project, we have identified a multigene family of at least 26 members encoding the t-SNARE syntaxin (PtSyx) that can be grouped into 15 subfamilies. Paramecium syntaxins match the classical build-up of syntaxins, being 'tail-anchored' membrane proteins with an N-terminal cytoplasmic domain and a membrane-bound single C-terminal hydrophobic domain. The membrane anchor is preceded by a conserved SNARE domain of approximately 60 amino acids that is supposed to participate in SNARE complex assembly. In a phylogenetic analysis, most of the Paramecium syntaxin genes were found to cluster in groups together with those from other organisms in a pathway-specific manner, allowing an assignment to different compartments in a homology-dependent way. However, some of them seem to have no counterparts in metazoans. In another approach, we fused one representative member of each of the syntaxin isoforms to green fluorescent protein and assessed the in vivo localization, which was further supported by immunolocalization of some syntaxins. This allowed us to assign syntaxins to all important trafficking pathways in Paramecium.     

Temperature effects on Korean entomopathogenic nematodes, Steinernema glaseri and S. longicaudum, and their symbiotic bacteria

We investigated the temperature effects on the virulence, development, reproduction, and motility of two Korean isolates of entomopathogenic nematodes, Steinernema glaseri Dongrae strain and S. longicaudum Nonsan strain. In addition, we studied the growth and virulence of their respective symbiotic bacterium, Xenorhabdus poinarii for S. glaseri and Xenorhabdus sp. for S. longicaudum, in an insect host at different temperatures. Insects infected with the nematode-bacterium complex or the symbiotic bacterium was placed at 13 degrees C, 18 degrees C, 24 degrees C, 30 degrees C, or 35 degrees C in the dark and the various parameters were monitored. Both nematode species caused mortality at all temperatures tested, with higher mortalities occurring at temperatures between 24 degrees C and 30 degrees C. However, S. longicaudum was better adapted to cold temperatures and caused higher mortality at 18 degrees C than S. glaseri. Both nematode species developed to adult at all temperatures, but no progeny production occurred at 13 degrees C or 35 degrees C. For S. glaseri, nematode progeny production was best at inocula levels above 20 infective juveniles/host at 24 degrees C and 30 degrees C, but for S. longicaudum, progeny production was generally better at 24 degrees C. Steinernema glaseri showed the greatest motility at 30 degrees C, whereas S. longicaudum showed good motility at 24 degrees C and 30 degrees C. Both bacterial species grew at all tested temperatures, but Xenorhabdus sp. was more virulent at low temperatures (13 degrees C and 18 degrees C) than X poinarii.     

Embryonic stem cells as a cellular model for neuroectodermal commitment and skin formation

Embryonic stem (ES) cells can be differentiated into many cell types in vitro, thus providing a potential unlimited supply of cells for cognitive in vitro studies and cell-based therapy. We recently reported the efficient derivation of ectodermal and epidermal cells from murine ES cells. These differentiated ES cells were able to form, in culture, a multilayered epidermis coupled with an underlying dermal compartment, similar to native skin. We clarified the function of BMP-4 in the binary neuroectodermal choice by stimulating sox-1(+) neural precursors to undergo specific apoptosis while inducing epidermal differentiation through DeltaNp63 gene activation. We further demonstrated that DeltaNp63 enhances ES-derived ectodermal cell proliferation and is necessary for epidermal commitment. This unique cellular model further provides a powerful tool for identifying the molecular mechanisms controlling normal skin development and for investigating p63-ectodermal dysplasia human congenital pathologies.     

Crystal structures of the pilus retraction motor PilT suggest large domain movements and subunit cooperation drive motility

PilT is a hexameric ATPase required for bacterial type IV pilus retraction and surface motility. Crystal structures of ADP- and ATP-bound Aquifex aeolicus PilT at 2.8 and 3.2 A resolution show N-terminal PAS-like and C-terminal RecA-like ATPase domains followed by a set of short C-terminal helices. The hexamer is formed by extensive polar subunit interactions between the ATPase core of one monomer and the N-terminal domain of the next. An additional structure captures a nonsymmetric PilT hexamer in which approach of invariant arginines from two subunits to the bound nucleotide forms an enzymatically competent active site. A panel of pilT mutations highlights the importance of the arginines, the PAS-like domain, the polar subunit interface, and the C-terminal helices for retraction. We present a model for ATP binding leading to dramatic PilT domain motions, engagement of the arginine wire, and subunit communication in this hexameric motor. Our conclusions apply to the entire type II/IV secretion ATPase family.     

Flowering phenology and survival of two annual plants <Emphasis Type="Italic">Impatiens noli-tangere</Emphasis> and <Emphasis Type="Italic">Persicaria thunbergii</Emphasis> co-occurring in streamside environments

Flowering phenology of cleistogamous annual plants Impatiens noli-tangere and Persicaria thunbergii was studied with reference to their mortality in streamside environments in northern Japan. I. noli-tangere produced flower-buds and flowers from late June to October, whereas P. thunbergii did so from mid-August to October. Both species were often killed by rapid current and submergence after heavy rainfall, which was unpredictable in the timing and intensity. P. thunbergii was more tolerant to rapid current and submergence than I. noli-tangere. Some individuals of I. noli-tangere were killed by herbivory from sawfly larvae in summer, but P. thunbergii individuals were seldom killed by herbivory. In consequence, the percentage of individuals that survived until the end of September was higher in P. thunbergii than in I. noli-tangere. These situations are assumed to select a long reproductive life in I. noli-tangere and autumn flowering in P. thunbergii.