Embryogenic callus formation,growth and regeneration in callus and suspension cultures of Miscanthus x ogiformis Honda Giganteus' as affected by proline

The effects of proline additions to culture systems of Miscanthus x ogiformis Honda Giganteus' were investigated. Proline was added in concentrations of 0, 12.5, 25, 50, 100 or 300 mM to the callus induction and suspension culture media containing either Murashige and Skoog or N6 basal salts and 22.6 μM 2,4-dichlorophenoxyacetic acid. Shoot apices and leaves from in vitro-propagated shoots, and immature inflorescences from greenhouse-grown plants were used as explants for callus induction and formation. Suspension cultures initiated from embryogenic callus of immature inflorescences were used to test the effect of proline in suspension cultures. The proline additions affected the formation of embryogenic callus and the growth of suspension cultures. Improvements depended on the proline concentration and the basal salts of the medium. Addition of 12.5 to 50 mM proline to callus induction medium with Murashige and Skoog salts increased embryogenic callus formation on shoot apices and leaf explants while proline had no effect on embryogenic callus formation in medium with N6 salts. Increased growth with increasing proline concentration was obtained in suspension aggregates grown in medium with N6 salts, whereas proline only increased growth of suspension aggregates grown in medium with Murashige and Skoog salts at concentrations of 12.5 or 25 mM. A stimulating effect of proline on plant regeneration was observed in short-term cultures of callus as well as in long-term cultures of suspension aggregates. An optimum proline concentration for plant regeneration was found at 12.5 mM. This revised version was published online in June 2006 with corrections to the Cover Date.     

Conditions affecting primary cell cultures of functional adult rat hepatocytes

Summary Primary monolayer cell cultures of adult rat hepatocytes underwent change in morphology and substantial cell loss between 1 and 3 days postinoculation. Dexamethasone-supplementation (1μM) of the culture medium maintained the polygonal epithelial morphology of the hepatocytes and increased longevity such that over 80% of the cells survived for 3 days and at least 30% for 8 or 9 days. This enhancement of survival was obtained up to 48 hr postinoculation, but the earlier the time of dexamethasone supplementation the greater the effect. Removal of dexamethasone resulted in a decrease in longevity. The positive effect of dexamethasone on longevity was observed following dexamethasone replacement of insulin in supplemented cultures, but the combination of insulin and dexamethasone resulted in poorer survival than with dexamethasone alone. The results are interpreted to indicate that dexamethasone provided a requirement of the in vitro environment for survival and suggest that elaboration of a complex medium is required to maintain hepatocytes in culture. This study was supported by an Alexander Ralston Peacock Memorial Grant for Cancer Research (No. BC-133A) from the American Cancer Society.     

Phenolsulfonphthalein (phenol red) metabolism in primary monolayer cultures of adult rat hepatocytes

Summary The sulfonic acid dye, phenolsulfonphthalein (PSP or phenol red), has been incorporated as a pH indicator in many tissue culture media formulations since the emergence of tissue culture methodologies. The present study was designed to examine the pathway, time course, and degree of metabolism of this anionic dye in monolayer cultures of adult rat hepatocytes. Thin layer chromatographic studies coupled with β-glucuronidase studies show that glucuronidation is the major metabolic pathway for PSP in vitro. About 20% of the dye is metabolized in the first 24 h, but this functional activity is decreased by approximately half at 48 h, and even further at 72 h of culture. This metabolic activity was not affected by continuous exposure to the dye. The effect of PSP concentration on its rate of metabolism by the adult rat hepatocyte in culture seemed to be biphasic, and at concentrations of less than 100μM there was indication of a saturable process. Although PSP seemed not to be toxic to hepatocyte cultures, it is partially metabolized by these cells (as opposed to no observed metabolism in human fibroblasts or HeLa cells). Therefore, its incorporation into tissue culture media formulations for use in hepatocyte cultures should be avoided, especially when studying the mechanism(s) of glucuronidation or metabolic pathways thought to be affected by this anionic dye. This study was supported in part by NIH Grants HL-11945-11 and 1 R01 AM 26520-01A1.     

Functional response of a parasitoid Ganaspidium utilis （Hymenoptera： Eucoilidae） on the leafminer Liriomyza trifolii （Diptera： Agromyzidae）

Functional response of a solitary, larval-pupal endoparasitoid of Liriomyza leafminers, Ganaspidium utilis Beardsley, was estimated on Liriomyza trifolii Burgess at three temperatures （17℃, 25℃, 29℃ ） and host densities. A type Ⅱ random parasitoid equation （RPE） was used to estimate instantaneous search rate and handling time. The instantaneous search rate increased as temperature increased. All of the RPE regressions obtained for functional response of G. utilis at different temperatures were significant （P〈0.01）. The slope of RPE regression lines was lower across the temperatures. At 29±2℃, the maximum number of larvae parasitized was 7.8 per day. It decreased to 7.2 larvae parasitized at 25±2℃. At 17±2℃, no significant increment of parasitization was observed due to the host density increments. The estimated handling time was lowest at 17±2℃ and highest at 25 ± 2℃, respectively. The ability of G. utilis to find and parasitize L. trifolii over a wide range of temperatures makes them a good candidate for biological control of Liriomyza leafminers.     

LSCHL4 from Japonica Cultivar,Which Is Allelic to NAL 1, Increases Yield of Indica Super Rice 93-11

The basic premise of high yield in rice is to improve leaf photosynthetic efficiency and coordinate the sourcesink relationship in rice plants. Quantitative trait loci （QTLs） related to morphological traits and chlorophyll content of rice leaves were detected at the stages of heading to maturity, and a major QTL （qLSCHL4） related to flag leaf shape and chlorophyll content was detected at both stages in recombinant inbred lines constructed using the indica rice cultivar 93-11 and the japonica rice cultivar Nipponbare. Map-based cloning and expression analysis showed that LSCHL4 is allelic to NAL1, a gene previously reported in narrow leaf mutant of rice. Overexpression lines transformed with vector carrying LSCHL4 from Nipponbare and a near-isogenic line of 93-11 （NIL-9311） had significantly increased leaf chlorophyll content, enlarged flag leaf size, and improved panicle type. The average yield of NIL-9311 was 18.70% higher than that of 93-11. These results indicate that LSCHL4 had a pleiotropic function. Exploring and pyramiding more high-yield alleles resem- bling LSCHL4 for super rice breeding provides an effective way to achieve new breakthroughs in raising rice yield and generate new ideas for solving the problem of global food safety.     

The effects of cell density, attachment substratum and dexamethasone on spontaneous apoptosis of rat hepatocytes in primary culture

Summary The rates of spontaneous cell detachment, cell viability, and apoptosis in primary cultures of rat hepatocytes plated at high and low density were compared. Apoptosis was frequent in detached cells, and the rates of cell detachment and apoptosis were greater in high-density than in low-density cultures. Among attached cells, more cells had condensed or fragmented nuclei in high-density than in low-density cultures. Further, ladder-like DNA fragmentation was not seen in low-cell-density cultures but was clearly evident in high-density cultures. Bax was more highly expressed in cells cultured at high density, and on collagen vs. matrigel, whereas changes of Bcl-2 and Fas expression observed in culture appeared unrelated to the rate of apoptosis. The rate of hepatocyte apoptosis appeared to be identical in low-density cultures on collagen 1 and matrigel, but when cells were cultured at high density, matrigel suppressed apoptosis by more than 50% at 36 h. In hepatocytes cultured on collagen 1, dexamethasone (0.1 μM) suppressed apoptosis in both low- and high-density cultures; higher doses had no further effects. In high density cultures, aurintricarboxylic acid (10 μM) suppressed apoptosis and this improved cell attachment at 48 h. It is concluded that cell viability in primary cultures of rat hepatocytes grown on collagen I is dependent on optimal culture density and that the cell population is regulated, at least in part, by apoptosis. Corticosteroids suppress spontaneous apoptosis of cultured hepatocytes in a non-dose-dependent manner, whereas matrigel abolishes apoptosis induced by increasing cell density. Bax may be an important protein in the cell density and cell matrix-dependent regulation of apoptosis in cultured hepatocytes.     

Sterol metabolism and oral epithelial cell growth

Summary Previous studies have demonstrated that as the density of cultured oral epithelial cells increases, there is a concomitant increase in phospholipids and cholesterol ester synthesis and a decrease in that of cholesterol and sterol precursors. Other studies have suggested that the effects of exogenous cholesterol sulfate may be similar to growth responses and influence metabolic steps related to cell density. To further examine this possibility, in the present study lipid synthesis was monitored in hamster cheek pouch epithelial cells in cultures established at different cells densities and in the presence of varying amounts of exogenous cholesterol sulfate. Cell [¹⁴C]acetate incorporation into lipids was measured in cultures established at four densities ranging from very subconfluent to very dense (postconfluent) in two media, Dulbecco’s modified Eagle’s medium (DMEM) with 5% fetal bovine serum and KSFM, a non-serum containing keratinocyte medium. Results indicated that the relative proportion of radiolabel incorporated into different lipid classes changed with cell density. In DMEM, the percentage of radiolabel incorporated into total phospholipids and fatty acids increased significantly with increasing cell density whereas percent incorporation into cholesterol, sterol precursors, and cholesterol esters significantly decreased. In KSFM cultures, proportionate phospholipids labeling was significantly increased in more dense cultures whereas cholesterol and cholesterol esters labeling was significantly decreased. In subconfluent and confluent cultures exposed to 10 or 25μM cholesterol sulfate, the relative proportions of phospholipid labeling also increased significantly compared to dimethyl sulfoxide (solvent) controls, whereas sterol precursors, fatty acids, and cholesterol esters labeling was signifcantly decreased. These results indicate that cholesterol sulfate can affect cellular lipid synthesis in a manner similar to that which occurs with increasing cell density, and strengthen the hypothesis that cholesterol sulfate may regulate lipid metabolic pathways related to growth and differentiation.     

Production protocol for and storage efficacy of an anthocorid predator Cardiastethus exiguus Poppius

Cardiastethus exiguus Poppius is an indigenous anthocorid predator of eggs and neonates of the notorious pest,coconut black-headed caterpillar Opisina arenosella Walker in India.At the National Bureau of Agriculturally Important Insects(Indian Council of Agricultural Research),Bangalore,India,a simple mass production protocol was developed for multiplying C.exiguus using UV-irradiated eggs of alternate laboratory host Corcyra cephalonica Stainton.Field evaluation of the predator in the states of Kerala and Karnataka indicated that this predator could bring about a significant reduction in the pest population.Subsequently,the need was felt to investigate the storage efficacy of the eggs and adults of C.exiguus so that sufficient numbers could be accumulated and transportation of the predator could be planned for field releases.Low temperature storage studies indicated that C.exiguus eggs can be safely stored for up to 5 days at 10℃ and 10 days at 15℃ and incubation period could be staggered for up to 10 and 13 days,respectively.The longevity of the C.exiguus adults was significantly reduced due to low temperature storage.However,for adult females,a storage temperature of 15℃ for 15 days could be recommended as they could live for a more than a month after removal from storage and their progeny production was comparable to that of the control adults.     

Short-term in vitro storage of t Miscanthus x ogiformis Honda 'Giganteus' as affected by medium composition,temperature, and photon flux density

Miscanthus x ogiformis Honda 'Giganteus' shoot cultures were stored in vitro on proliferation or rooting medium for up to 27 weeks at temperatures of 8, 12, 16, or 20 °C and photosynthetic photon flux densities of 5, 10, or 20 μmol m⁻² s⁻¹. Plants survived storage much better on rooting medium than on proliferation medium. Plants stored on rooting medium for 1 week survived well when survival was assessed immediately after storage or after 14 days of acclimatization, but had the lowest survival 28 days after transplantation. With increasing storage period on rooting medium increasing survival was found 28 days after transplantation. This was probably a result of the development of rhizomes and/or roots during storage. Best survival was observed at 20 μmol m⁻² s⁻¹ and a temperature of 8-16 °C. Increasing the temperature to 25 °C during the last week of storage improved survival considerably. Root formation was slow at 8 °C, but after 27 weeks of storage the rooting percentage was the same at all storage temperatures. An increasing number of shoots per plant 28 days after transplantation was found with increasing PPFD during storage.Miscanthus shoot cultures can be stored in vitro for at least 27 weeks with limited losses when stored on rooting medium at 20 μmol m⁻² s⁻¹, a temperature of 16 °C, and given a 1-week end-of-storage treatment of 25 °C. This revised version was published online in June 2006 with corrections to the Cover Date.     

Heat sensitivity of eggs attributes to the reduction in Agasicles hygrophila population

Agasicles hygrophila has been introduced worldwide as a control agent for the invasive weed Alternanthera philoxeroides.However,global warming has potential impact on its controlling efficacy.The aim of this research was to explore the primary factors responsible for the greatly reduced A.hygrophila population in hot summers.To imitate the temperature conditions in summers,different developmental stages of hygrophila were treated with high temperatures from 32.5℃ to 45℃ for 1-5 h.Based on the survival rate,the heat tolerance of each developmental stage was ranked from lowest to highest as follows:egg,1st,2nd,3rd instar larva,adult and pupa.Eggs showed the lowest heat tolerance with 37.5℃ as the critical temperature affecting larval hatching.Heat treatment of the A.hygrophila eggs at 37.5℃ for 1 h decreased the hatch rate to 24%.Our results indicated that when compared with the control at 25℃,1 h treatment at 37.5℃ prolonged the duration of the egg stage,shortened the duration of oviposition and total longevity,and changed the reproductive pattern of A.hygrophila.The net reproductive rate,intrinsic rate and finite rate were all significantly reduced.The results suggest that low heat tolerance of the eggs was the major factor responsible for the reduction of A.hygrophila populations,and the key temperature was 37.5°C.Therefore,appropriate measures should be taken to protect eggs in order to maintain the efficacy of A.hygrophila in the biological control of A.philoxeroides in hot summers.     

Recent advances in the study of Kaposi's sarcoma-associated herpesvirus replication and pathogenesis

It has now been over twenty years since a novel herpesviral genome was identified in Kaposi's sarcoma biopsies. Since then, the cumulative research effort by molecular biologists, virologists, clinicians, and epidemiologists alike has led to the extensive characterization of this tumor virus, Kaposi's sarcoma-associated herpesvirus(KSHV; also known as human herpesvirus 8(HHV-8)), and its associated diseases. Here we review the current knowledge of KSHV biology and pathogenesis, with a particular emphasis on new and exciting advances in the field of epigenetics. We also discuss the development and practicality of various cell culture and animal model systems to study KSHV replication and pathogenesis.     

A Unique Protease Cleavage Site Predicted in the Spike Protein of the Novel Pneumonia Coronavirus (2019-nCoV) Potentially Related to Viral Transmissibility

正Dear Editor,In December 2019, a novel human coronavirus caused an epidemic of severe pneumonia(Coronavirus Disease 2019,COVID-19) in Wuhan, Hubei, China(Wu et al. 2020; Zhu et al. 2020). So far, this virus has spread to all areas of China and even to other countries. The epidemic has caused 67,102 confirmed infections with 1526 fatal cases     

Curcuminoids as inhibitors of thioredoxin reductase: A receptor based pharmacophore study with distance mapping of the active site

Curcumin is the yellow pigment of turmeric that interacts irreversibly forming an adduct with thioredoxin reductase (TrxR), an enzyme responsible for redox control of cell and defence against oxidative stress. Docking at both the active sites of TrxR was performed to compare the potency of three naturally occurring curcuminoids, namely curcumin, demethoxy curcumin and bis-demethoxy curcumin. Results show that active sites of TrxR occur at the junction of E and F chains. Volume and area of both cavities is predicted. It has been concluded by distance mapping of the most active conformations that Se atom of catalytic residue SeCYS498, is at a distance of 3.56 from C13 of demethoxy curcumin at the E chain active site, whereas C13 carbon atom forms adduct with Se atom of SeCys 498. We report that at least one methoxy group in curcuminoids is necessary for interation with catalytic residues of thioredoxin. Pharmacophore of both active sites of the TrxR receptor for curcumin and demethoxy curcumin molecules has been drawn and proposed for design and synthesis of most probable potent antiproliferative synthetic drugs.     

The structure, reproduction and taxonomy of Vidalia and Osmundaria (Rhodophyta, Rhodomelaceae)

Comprises species occurring mostly in subtidal habitats in tropical, subtropical and warm-temperate areas of the world. An analysis of the type species, V. spiralis (Sonder) Lamouroux ex J. Agardh, a species from Australia, establishes basic characters for distinguishing species in the genus. These characters are (1) branching patterns of thalli, (2) flat blades that may be spiralled on their axis, (3) width of the blade, (4) primary or secondary derivation of sterile and fertile branchlets and (5) position of sterile and fertile branchlets on the thalli. Application of the latter two characters provides an important basic method for separation of species into three major groups. Osmundaria , a genus known only in southern Australia, was studied in relation to Vidalia , and its separation from the Vidalia assemblage is not accepted. Species of Vidalia therefore are transferred to the older genus name, Osmundaria. Two new species, Osmundaria papenfussii and Osmundaria oliveae are described from Natal. Confusion in the usage of the epithet, Vidalia fimbriala Brown ex Turner has been clarified, and Vidalia gregaria Falkenberg, described as an epiphyte on Osmundaria pro/ifera Lamouroux, is revealed to be young branches of the host, Osmundaria prolifera.     

New or rare chromosome counts in Artemisia L. (Asteraceae, Anthemideae) and related genera from Kazakhstan

Fifteen chromosome counts of six Artemisia taxa and one species of each of the genera Brachanthemum, Hippolytia, Kaschgaria, Lepidolopsis and Turaniphytum are reported from Kazakhstan. Three of them are new reports, two are not consistent with previous counts and the remainder are confirmations of very scarce (one to four) earlier records. All the populations studied have the same basic chromosome number, x = 9, with ploidy levels ranging from 2x to 6x. Some correlations between ploidy level, morphological characters and distribution are noted.