Controlled Growth and the Maintenance of Human Pluripotent Stem Cells by Cultivation with Defined Medium on Extracellular Matrix-Coated Micropatterned Dishes

Here, we introduce a new serum-free defined medium (SPM) that supports the cultivation of human pluripotent stem cells (hPSCs) on recombinant human vitronectin-N (rhVNT-N)-coated dishes after seeding with either cell clumps or single cells. With this system, there was no need for an intervening sequential adaptation process after moving hPSCs from feeder layer-dependent conditions. We also introduce a micropatterned dish that was coated with extracellular matrix by photolithographic technology. This procedure allowed the cultivation of hPSCs on 199 individual rhVNT-N-coated small round spots (1 mm in diameter) on each 35-mm polystyrene dish (termed “patterned culture”), permitting the simultaneous formation of 199 uniform high-density small-sized colonies. This culture system supported controlled cell growth and maintenance of undifferentiated hPSCs better than dishes in which the entire surface was coated with rhVNT-N (termed “non-patterned cultures”). Non-patterned cultures produced variable, unrestricted cell proliferation with non-uniform cell growth and uneven densities in which we observed downregulated expression of some self-renewal-related markers. Comparative flow cytometric studies of the expression of pluripotency-related molecules SSEA-3 and TRA-1-60 in hPSCs from non-patterned cultures and patterned cultures supported this concept. Patterned cultures of hPSCs allowed sequential visual inspection of every hPSC colony, giving an address and number in patterned culture dishes. Several spots could be sampled for quality control tests of production batches, thereby permitting the monitoring of hPSCs in a single culture dish. Our new patterned culture system utilizing photolithography provides a robust, reproducible and controllable cell culture system and demonstrates technological advantages for the mass production of hPSCs with process quality control.     

Regulation of vascular smooth muscle cells by micropatterning

Vascular smooth muscle cells (SMCs) undergo morphological and phenotypic changes when cultured in vitro. To investigate whether SMC morphology regulates SMC functions, bovine aortic SMCs were grown on micropatterned collagen strips (50-, 30-, and 20-microm wide). The cell shape index and proliferation rate of SMCs on 30- and 20-microm strips were significantly lower than those on non-patterned collagen (control), and the spreading area was decreased only for cells patterned on the 20-microm strips, suggesting that SMC proliferation is dependent on cell shape index. The formation of actin stress fibers and the expression of alpha-actin were decreased in SMCs on the 20- and 30-microm collagen strips. SMCs cultured on micropatterned biomaterial poly-(D,L-lactide-co-glycolide) (PLGA) with 30-microm wide grooves also showed lower proliferation rate and less stress fibers than SMCs on non-patterned PLGA. Our findings suggest that micropatterned matrix proteins and topography can be used to control SMC morphology and that elongated cell morphology decreases SMC proliferation but is not sufficient to promote contractile phenotype.     

Fabrication and characterization of a multiwell array SERS chip with biological applications

Uniform, large surface area substrates for surface-enhanced Raman spectroscopy (SERS) are fabricated by oblique angle deposition. The SERS-active substrates are patterned by a polymer-molding technique to provide a uniform array for high throughput biosensing and multiplexing. Using a conventional SERS-active molecule, 1,2-di(4-pyridyl)ethylene (BPE) ≥98%, we show that this device provides a uniform Raman signal enhancement from well to well with a detection limit of at least 10⁻⁸ M of the BPE solution or 10⁻¹⁸ mol of BPE. The SERS intensity is also demonstrated to vary logarithmically with the log of BPE concentration and the apparent sensitivity of the patterned substrate is compared to previous reports from our group on non-patterned substrates. Avian influenza is analyzed to demonstrate the utility of SERS multiwell patterned substrates for biosensing. The spectra acquired from patterned substrates show better reproducibility and less variation compared to the unpatterned substrates according to multivariate analysis. Our results highlight potential advantages of the patterned substrate.     

The startle responses of blue jays to Catocala (Lepidoptera: Noctuidae) prey models

Artificial moth models were presented to caged blue jays, Cyanocitta cristata, in order to investigate the effectiveness of deimatic displays in underwing (Catocala: Noctuidae) moths. The models had flexible, patterned ‘hindwings’ which were concealed behind cardboard ‘forewings’ until the birds removed them from a presentation board. Jays which had been trained on models with grey hindwings exhibited a startle response when they were exposed to Catocala-patterned hindwings. In contrast to this, subjects trained on Catocala models did not startle to a novel grey hindwing. The startle response to Catocala patterns lasted over several days until birds habituated to the models. When the jays had habituated to one Catocala hindwing pattern, a novel Catocala pattern always elicited a startle response. Familiar Catocala hindwing patterns which appeared in an anomalous context (i.e., associated with a different forewing pattern) also evoked a startle response from these birds. Novelty, oddity, conspicuousness, and anomaly are considered as possible stimulus characteristics which trigger the startle response.     

Patterns and niches in diachronic word formation: the fate of the suffix -<Emphasis Type="SmallCaps">men</Emphasis> from Latin to Romance

The Latin suffix -men of words like foramen, nutrimen, albumen, etc. has left traces in all Romance idioms, but the outcome has been quite different in each of them. The present paper describes in some detail the distribution and the meaning of the successors of -men in each of the Romance idioms and tries to uncover the mechanisms behind the fragmentation process. Some of the changes already took place in Latin, while others occurred in Romance times. It is argued that in the absence of lexical continuities between Latin and Romance a close attention to word-formation patterns and semantic niches, as well as a close comparison of similar developments in Romance idioms can be helpful in reconstruction.     

Differential gene expression analysis of ‘Granny Smith’ apple (Malus domestica Borkh.) during fruit skin coloration

‘Granny Smith’ apples growing under normal sunlight develop green skin, whereas the peel turns red due to anthocyanin accumulation after the removal of a bagging treatment. Two anthocyanins, Cyanidin 3-O-galactoside (cy3-gal) and Cyanidin 3-O-arabinoside (cy3-ara), were detected in the red ‘Granny Smith’ apple peels, and cy3-gal was determined to be chiefly responsible for the red color. The content of cy3-gal was more than 98% of the total anthocyanin in the red ‘Granny Smith’ peels. To better understand the molecular basis of anthocyanin biosynthesis in ‘Granny Smith’ apples, we performed a quantitative real-time PCR (qRT-PCR) analysis of anthocyanin biosynthetic genes (MdCHS, MdF3H, MdDFR, MdANS, MdUFGT, and MdMYB1). Our results indicate that the expression of these genes (except MdCHS) was associated with increased anthocyanin accumulation in the skin of ‘Granny Smith’ apples. Four selected genes obtained from the ‘Granny Smith’ skin cDNA library, phytoene synthase (PSY), WD40 repeat protein, polygalacturonase (PG), and galactosidase (GAL), were also confirmed by qRT-PCR. We found that these genes were differently expressed during ‘Granny Smith’ apple skin coloration, suggesting that they are directly or indirectly involved in pigment accumulation. In conclusion, anthocyanin biosynthesis in ‘Granny Smith’ apples is the result of interactions between multiple enzymes in the anthocyanin biosynthesis pathway, and the coloring mechanism of ‘Granny Smith’ apples may be similar to that of red-skinned cultivars.     

Biology of the Two-Spotted Spider Mite on Strawberry Plants

The two-spotted spider mite, Tetranychus urticae Koch, is commonly found on strawberry crops (Fragaria x ananassa). Strawberry plants have defensive mechanisms, which in turn influence the behavior of herbivores. The oviposition and development of the two-spotted spider mite were evaluated on the leaf disks of the cultivars ‘Aromas,’ ‘Camarosa,’ ‘Camino Real,’ ‘Diamante,’ ‘Diamante 10,’ ‘Diamante 50,’ ‘Festival,’ and ‘Seascape.’ It was observed that on cultivars such as ‘Aromas,’ ‘Camarosa,’ and ‘Seascape,’ immature survivorship was higher, but no difference was found during the developmental period from egg to adult of T. urticae. The immature development time was also longer on ‘Camarosa.’ Females laid more eggs on ‘Seascape’ (8.4 eggs/day), and the least on ‘Camarosa’ (1.0 egg/day). Mortality was higher at the larval stage and reached more than 50% in three cultivars ‘Camarosa,’ ‘Diamante,’ and ‘Seascape.’ Thus, the cultivars ‘Camarosa,’ ‘Diamante,’ and ‘Seascape’ were the ones that mostly affected the survival, development, and reproduction of T. urticae.     

Identification and analysis of cold stress-inducible genes in Korean rapeseed varieties

The rapeseed plant (Brassica napus) is one of the top five oil-producing crops in the world. However, its potential for global cultivation is limited due to its sensitivity to cold. We selected five varieties of rapeseed popular in Korea and analyzed their sensitivity to cold stress. Three — ‘Naehan’, ‘Sandongchae’, and ‘Yudal’ — proved more tolerant than the other two — ‘Youngsan’ and ‘Tammi’. Because rapeseed is in the same family (Brassicacae) as Arabidopsis, one would expect that both share similar genes, with protein coding sequences that show >86% homology. We used a cDNA microarray enriched with Arabidopsis cold stress-related genes to examine the molecular background for differences in sensitivity among rapeseed varieties. Their hierarchical clustering patterns for expression were highly correlated with their cold-sensitive phenotypes. For example, expression patterns were similar between ‘Naehan’ and ‘Sandongchae’ as well as between ‘Yudal’ and our Arabidopsis samples. Using co-expression gene sets developed in this laboratory, we further analyzed expression patterns for three cultivars. Three distinct gene sets were commonly enriched in ‘Naehan’ against ‘Youngsan’ and ‘Tammi’, indicating that they could be the core gene sets that confer a high degree of cold tolerance in ‘Naehan’. Knockout plants with down-regulated expression of GSTF2, TRXh5, LTI30, and LACS4 were highly cold-sensitive whereas overexpression of those four genes led to a reversal of this sensitivity. In fact, overexpression of GSTF2 and LTI30 enhanced tolerance, with plants that over-expressed GSTF2 having extremely high tolerance that enabled them to acclimate almost completely to freezing conditions. By comparison, only about 40% of the wild type demonstrated a similar acclimation. Therefore, the genes described here could be useful in furthering our knowledge about other stress responses in rapeseed, thereby facilitating the production of cold-tolerant plants.     

Intraspecific variation in alkaloid profile of four lupine species with implications for the pea aphid probing behaviour

The alkaloid profile of white lupine Lupinus albus L. cv. ‘Boros’ and ‘Butan’, narrow-leaf lupine L. angustifolius L. cv. ‘Bojar’, ‘Graf’, ‘Karo’, ‘Mirela’, and ‘Sonet’, yellow lupine L. luteus L. cv. ‘Dukat’, ‘Parys’, ‘Perkoz’, and ‘Talar’, and wild species big-leaf lupine L. polyphyllus Lindl. was studied. The pea aphid Acyrthosiphon pisum Harris probing behaviour was monitored using the electrical penetration graph (EPG) technique.Four types of feeding behaviour were recognized. First, on highly acceptable L. luteus cv. ‘Dukat’, the total and mean duration of probing, time to reach phloem phase, and the duration of the first phloem phase were comparable to those in aphids on control plant Pisum sativum. Second, on partially acceptable L. luteus cv. ‘Talar’, pathway activities were slightly impeded, and the probes were more numerous and slightly shorter than on control plants. The phloem phase occurred sporadically, and feeding was terminated early after a brief period of ingestion. Third, on unpalatable but acceptable L. angustifolius cv. ‘Bojar’ and ‘Sonet’, and L. luteus cv. ‘Parys’ and ‘Perkoz’, the probes were numerous and usually epidermal. The phloem phase occurred rarely, and when it did, it was short and consisted mainly of watery salivation. Finally, on unacceptable L. albus cv. ‘Boros’ and ‘Butan’, L. angustifolius cv. ‘Graf’, ‘Karo’, ‘Mirela’, and L. polyphyllus, total probing time and individual probes were short and phloem phase did not occur.Eighteen alkaloids were identified: one piperidine alkaloid (ammodendrine), one indole (gramine), ten quinolizidine alkaloids (one tricyclic and nine tetracyclic compounds), and six esters. All lupine varieties that contained lupanine, its derivatives and especially their esters appeared to be unacceptable to the pea aphid, independent of the total concentration of any specific lupanine alkaloid. In contrast, sparteine and its derivatives did not seem to affect aphid probing significantly.     

Characterization of self-compatibility in sweet cherry varieties by crossing experiments and molecular genetic analysis

Self-compatibility is a major breeding objective in sweet cherry. The identification and characterization of new sources of self-compatibility will be useful for breeding and research purposes. In this work, self-compatibility of four local Spanish sweet cherry varieties was investigated by crossing experiments and molecular genetic analysis of two self-incompatibility loci. Crossing experiments included self- and cross-pollinations in the laboratory followed by microscopic observation of pollen tube growth and fruit set assay in the field. After crossing experiments, two accessions, ‘Son Miró’ and ‘Talegal Ahín’, were self-compatible while the other two were self-incompatible. Inheritance of S-locus and microsatellite EMPaS02 (linked to self-compatibility, Sc) were investigated in self-pollination progeny of both self-compatible genotypes. Results indicate that self-compatibility in ‘Talegal Ahín’ is similar to self-compatibility described in sweet cherry ‘Cristobalina’ and may be caused by the same mutation. That is a pollen part mutation not linked to the S-locus but linked to microsatellite EMPaS02 in cherry LG3. In ‘Son Miró’ self-compatibility seems more complex, affecting pollen and style function, and probably involving more than one mutation not described previously in sweet cherry. Together with ‘Cristobalina’, the newly described self-compatible varieties ‘Son Miró’ and ‘Talegal Ahín’ confirm the existence of unique self-compatible plant material in local germplasm from Spain that should be conserved and characterized for its use in breeding and research.     

A SNP in the promoter region of the<Emphasis Type="Italic">VvmybA1</Emphasis> gene is responsible for differences in grape berry color between two related bud sports of grape

The VvmybA1 gene in grape (Vitis vinifera) plays a key role in the biosynthesis of anthocyanin. The grape cultivars, ‘Benitaka’ (red color) and ‘Brazil’ (black color) were the result of a bud mutation. ‘Benetika’ was derived from ‘Italia’ (green color) and ‘Brazil’ was developed from ‘Benetika’. Single sequence repeat (SSR) molecular marker analysis was performed in order to demonstrate that the three cultivars have a common pedigree. A sequence analysis of the promoter region and coding sequence of VvmybA1 revealed a base substitution between ‘Benitaka’ and ‘Brazil’ in the promoter region and a deletion of a large DNA fragment in the promoter region of ‘Italia’. Anthocyanin content and expression of the VvmybA1 and UFGT genes in ‘Brazil’ were higher than in ‘Benitaka’ and barely detectable in ‘Italia’. A transient expression system was used to introduce VvmybA1 driven by the three different promoters present in ‘Italia’, ‘Brazil’, and ‘Benitaka’ into somatic embryos of ‘Centennial Seedless’ (Vitis vinifera L.) by Agrobacterium-mediated transformation. This resulted in the production of red cells in the embryos transformed with the constructs of VvmybA1 from ‘Brazil’ and ‘Benitaka’ and no color production in embryos transformed with the VvmybA1 construct from ‘Italia’. In addition, the embryos transformed with the ‘Brazil’ construct had more red color than the embryos transformed with the ‘Benitaka’ construct. These results suggested that a SNP mutation in the promoter region of VvmybA1 in ‘Benitaka’ (red color) was responsible for the color change displayed by ‘Brazil’ (black color).     

‘Endong’, a rare variety of the farmed carrageenophyte Eucheuma denticulatum (Burman) Collins & Hervey from the Philippines

The variety described in this paper is rather a rare and interesting one. Locally called ‘endong’, this was discovered first in a farm in Tawi-Tawi, Sulu Archipelago, Philippines. During harvest, seaweed farmers used to mix this with ‘tambalang’ (Kappaphycus alvarezii (Doty) Doty ex Silva var. tambalang Doty, the ‘cottonii’ of commerce) because of the presumption of it producing the same carrageenan type as that of ‘tambalang’, viz., kappa-carrageenan. The two farmed varieties are similar in appearance. Complaints arose from seaweed processing plants when the resulting gelling properties (hard and brittle gel) expected of the kappa-carrageenan-producing ‘tambalang’ were not met. Because of its lax habit and long, smooth, and naked basal axes, ‘endong’ was initially thought to be related to ‘tambalang’ and differ from the farmed ‘spinosum’ (Eucheuma denticulatum (Burman) Collins & Hervey) whose typical or common form has shorter axes covered from base to apex with spinous determinate (ramuli) or indeterminate branchlets arranged irregularly and in whorls at predictable interval. However, a closer look at ‘endong’ shows that it also has spinous branchlets arranged in whorls, but confined to the apical portion of the branches. Moreover, its internal axial anatomy shows rhizoidal filaments forming a core. Both characters fit well the distinguishing taxonomic criteria established for E. denticulatum. Chemical characterization of its carrageenan and the determination of its phylogenetic affinities support our morphological findings. Hence, we report and describe ‘endong’ as a rare variety, and we name it E. denticulatum (Burman) Collins & Hervey var. endong Trono & Ganzon-Fortes var. nov. Tips to distinguish ‘endong’ from ‘tambalang’ are included in this paper to aid seaweed farmers in separating the two during harvest.     

Where Did Information Go? Reflections on the Logical Status of Information in a Cybernetic and Semiotic Perspective

This article explores the usefulness of interdisciplinarity as method of enquiry by proposing an investigation of the concept of information in the light of semiotics. This is because, as Kull, Deacon, Emmeche, Hoffmeyer and Stjernfelt state, information is an implicitly semiotic term (Biological Theory 4(2):167–173, 2009: 169), but the logical relation between semiosis and information has not been sufficiently clarified yet. Across the history of cybernetics, the concept of information undergoes an uneven development; that is, information is an ‘objective’ entity in first order cybernetics, and becomes a ‘subjective’ entity in second order cybernetics. This contradiction relegates the status of information to that of a ‘true’ or ‘false’ formal logic problem. The present study proposes that a solution to this contradiction can be found in Deely’s reconfiguration of Peirce’s ‘object’ (as found in his triadic model of semiosis) into ‘thing’ and ‘object’ (Deely 1981). This ontology allows one to argue that information is neither ‘true’ nor ‘false’, and to suggest that, when considered in light of its workability, information can be both true and false, and as such it constitutes an organism’s purely objective reality (Deely 2009b). It is stated that in the process of building such a reality, information is ‘motivated’ by environmental, physiological, emotional (including past feelings and expectations) constraints which are, in turn, framed by observership. Information is therefore found in the irreducible cybersemiotic process that links at once all these conditions and that is simultaneously constrained by them. The integration of cybernetics’ and semiotics’ understanding of information shows that history is the analytical principle that grants scientific rigour to interdisciplinary investigations. As such, in any attempt to clarify its epistemological stance (e.g. the semiotic aspect of information), it is argued that biosemiotics does not need only to acknowledge semiotics (as it does), but also cybernetics in its interdisciplinary heritage.     

A comparison of the geometric configurations adopted by radiation-induced chromatid interchanges in animals and plants

An analysis of radiation-induced chromatid interchanges in four animal and five plant species indicates that polarised (P) and X-type interchanges occur relatively more frequently in the plants, but non-polarised (N) and U type interchanges occur more frequently in the animals. An explanation of this difference, compatible with the observed geometric shapes of these interchanges and their differing propensity to incompleteness, is that; (a) in all the organisms examined, U- and X-type exchanges are formed during interphase between polarised chromosomes, i.e. they are notional ‘PU’ and ‘PX’ types, (b) the ratio ‘PU’ : ‘PX’ is similar in both animals and plants, (c) a proportion of these exchanges is established during interphase between looped and unlooped (and possibly terminal overlapping) segments, and this proportion is greater in animals; as a result of subsequent chromatin condensation these ‘PU’ and ‘PX’ exchanges may be converted to NU and NX types respectively, (d) ‘PX’ exchanges which are not converted by this mechanism, and NX exchanges which arise in late prophase as a result of this mechanism, are unstable and may be converted during prometaphase to the more stable NU and PU configurations respectively, (e) transformations (d) are relatively more frequent in the animal species because the flexural and tensional regidities of the chromosomes are greater than in the plant chromosomes.     

The odontode explosion: The origin of tooth‐like structures in vertebrates

Essentially we show recent data to shed new light on the thorny controversy of how teeth arose in evolution. Essentially we show (a) how teeth can form equally from any epithelium, be it endoderm, ectoderm or a combination of the two and (b) that the gene expression programs of oral versus pharyngeal teeth are remarkably similar. Classic theories suggest that (i) skin denticles evolved first and odontode‐inductive surface ectoderm merged inside the oral cavity to form teeth (the ‘outside‐in’ hypothesis) or that (ii) patterned odontodes evolved first from endoderm deep inside the pharyngeal cavity (the ‘inside‐out’ hypothesis). We propose a new perspective that views odontodes as structures sharing a deep molecular homology, united by sets of co‐expressed genes defining a competent thickened epithelium and a collaborative neural crest‐derived ectomesenchyme. Simply put, odontodes develop ‘inside and out’, wherever and whenever these co‐expressed gene sets signal to one another. Our perspective complements the classic theories and highlights an agenda for specific experimental manipulations in model and non‐model organisms.     

From d’Orbigny to the Devonian: some thoughts on the history of the stratotype concept

D’Orbigny’s stratigraphic contributions were enormous, both practical and theoretical. In his practice, d’Orbigny realised the central importance of using fossil faunas to correlate strata across different countries. His ‘succession chronologique des âges du monde’ (of 1852) recognised 5 periods and 27 stages (ignoring his ‘époque actuelle’). The great majority of the latter were from two Periods, Jurassic (stages 7 to 16) or Cretaceous (17 to 23). D’Orbigny described the palaeontological characteristics, and geographical extensions, of each. Some stages he based on countries he never visited and although he never specified ‘stratotype’ localities, he referred to some as ‘étalon’, ‘le mieux’ or ‘le plus beau’. These leads proved crucial when, in the mid 20th century, stratigraphers realised that definitions of chrono-stratal units must be attempted. The first attempts at typification simply followed zoology, where a type specimen tries to define a central position within the morphological variation of a species. In 1962, an early attempt was made to define Jurassic stages by three type horizons at different type localities (or stratotypes as they were now called) for each. A central horizon was chosen as ‘lectotype section’ and upper and lower boundaries for each were defined elsewhere. In France, it was preferred to investigate original intentions at named localities, through unit-stratotypes. These allowed some nationalistic pride, since France was the home of so many d’Orbignyan stages. 1980 saw the publication Les Étages français et leurs Stratotypes. However, British Silurian stratigraphers had realised that such zoologically based concepts would produce conflict, when any defined upper boundary stratotype conflicted with the defined lower boundary of a superjacent unit. They suggested, from 1962, to define only lower boundaries, via ‘golden spikes’, at basal-boundary stratotypes. This was the method chosen for the basal Devonian stratotype at Klonk, Czech Republic, defined in 1972, and a method adopted globally from 1986. However, this was when correlations were still largely based on fossils. The explosion of so many other stratigraphies since, based on magnetic or chemical records, events etc, has produced a first reaction against the idea that such stratotypes should be so defined, while a second, potentially more major claim, is that any ‘golden spike’ concept may no longer ‘glitter’, as competing stratotype concepts may be holding back stratigraphic progress.     

Resistance of Citrus and Related Genera to Diaphorina citri Kuwayama (Hemiptera: Liviidae)

The present study was developed to evaluate the resistance of the following genotypes of Citrus and related genera to this pest: ‘Pera,’ ‘Natal’, and ‘Washington Navel’ oranges (Citrus sinensis), ‘Marsh Seedless’ grapefruit (Citrus paradisi), hardy orange ‘Rubidoux’ (Poncirus trifoliata), kumquat (Fortunella margarita Swingle), citrumelo ‘Swingle’ (C. paradisi x P. trifoliata), and citrange ‘Troyer’ (P. trifoliata x C. sinensis). The experiments were performed in greenhouses with plants grafted onto ‘Rangpur’ lime (Citrus limonia) and placed individually in voile cages. The preference for oviposition in a no-choice test, and the effect of genotype were evaluated. The egg-adult cycle was monitored to determine the effect of genotype on the biology of the insect. Poncirus ‘Rubidoux’ was the least preferred genotype for oviposition; reduced number of eggs was also found to occur on citrange ‘Troyer’, and ‘Marsh Seedless’ was the genotype with the most eggs. No significant variation in the duration of the embryonic period was observed; however, a difference in the viability of eggs was found, with the lowest egg viabilities on ‘Swingle.’ Kumquat and ‘Marsh Seedless’ genotypes were correlated with increased durations of the nymphal phase, however, there was no difference in the survival of this phase. Fecundity of females on ‘Troyer’, ‘Swingle’, and kumquat was reduced. Considering all of the evaluated parameters, it was concluded that cultivars of sweet orange are the most susceptible genotypes to Diaphorina citri. Regarding oviposition, P. trifoliata ‘Rubidoux’ showed resistance of the antixenosis type.     

Effects of virus infection on plant growth,root development and phytohormone levels in in vitro-cultured pear plants

Virus infection is common in pear (Pyrus spp.) trees commercially cultivated in China. In this study, the growth speed, root development and phytohormone levels of virus-free and virus-infected pear cultures were comparatively investigated. Results showed that the co-infection of Apple stem grooving virus (ASGV) and Apple stem pitting virus (ASPV) significantly decreased the growth and proliferation of in vitro plants of P. communis cv. ‘Confenence’, but showed relatively less effect for P. pyrifolia cv. ‘Jinshui no. 2’, P. communis cv. ‘Red Clapp Favonite’ and P. sinkiangensis cv. ‘Korla’. Strong inhibition of ASGV infection to root development was observed for in vitro plants of ‘Jinshui no. 2’, but not for ‘Confenence’. Furthermore, during root induction, ASGV infection significantly increased CTKs/IAA ratio and decreased IAA/ABA ratio in the rooting region of in vitro plants of ‘Jinshui no. 2’. Nevertheless, for in vitro plants of ‘Confenence’, these values were rarely influenced by ASGV and ASPV infection. The result indicated that the changes of CTKs/IAA and IAA/ABA ratios might be responsible for rooting inhibition of in vitro plants of ‘Jinshui no. 2’.     

<Emphasis Type="Italic">Muscadinia rotundifolia</Emphasis> ‘Noble’ defense response to <Emphasis Type="Italic">Plasmopara viticola</Emphasis> inoculation by inducing phytohormone-mediated stilbene accumulation

Downy mildew (DM), one of the most devastating grape diseases worldwide, is caused by the biotrophic oomycete Plasmopara viticola (Pv). In general, grapevine responds to Pv infection with the accumulation of phytoalexins as part of the innate immune system, and diverse phytoalexins are induced on grapevines with different DM-resistance levels in response to Pv invasion. However, the regulation of phytoalexin biosynthesis during grapevine against Pv is still unclear. Herein, we detected stilbenes by UPLC-ESI-MS/MS and found that resveratrol was accumulated to higher level and earlier in the DM-immune Muscadinia rotundifolia ‘Noble’ than that in the DM-susceptible Vitis vinifera ‘Thompson Seedless’ after Pv inoculation. Additionally, a considerable amount of pterostilbene and ε-viniferin was found in ‘Noble’, while a little was detected in ‘Thompson Seedless’. Resveratrol was glycosylated into piceid both in ‘Noble’ and ‘Thompson Seedless’ after Pv inoculation. The qPCR analysis of gene expression indicated that the resveratrol-synthesis gene (STS) was induced by Pv inoculation earlier in ‘Noble’ than that in ‘Thompson Seedless’, while the pterostilbene-synthesis gene (ROMT) was induced in ‘Noble’ but not in ‘Thompson Seedless’ at all. The piceid-synthesis gene (GT) was generally up-regulated in both cultivars. Sequence analysis of STS, ROMT, and GT promoters revealed that they contained cis-regulatory elements responsive to phytohormones and pathogens. Following Pv inoculation, the level of SA, MeJA, and ABA was found to be consistently higher in ‘Noble’ than those in ‘Thompson Seedless’. The results of exogenous hormone elicitation further demonstrated that the accumulation of stilbenes was regulated by phytohormones. The earlier and higher accumulation of phytohormones and consequent induction of stilbene synthesis may play an important role in grapevine defense against downy mildew disease.