In vitro selection and field responses of somaclonal variant plants of rice cv PR113 for drought tolerance

Drought is the major environmental stress that limits rice productivity worldwide. In vitro somaclonal variation using different selection agents has been used for crop improvement. Here, rice plants of cv PR113 were selected in vitro on 30, 50 and 70 g L^-1 polyethylene glycol 6,000 (PEG). Callus growth, proliferation, calli volume (first and second culture) and plantlet regeneration (third culture) were found to be decreased upto a certain level to acquire tolerance to PEG-induced drought. From the field data, 30 g L^-1 PEG lines showed higher vegetative growth (plant height, tiller number, leaf number, shoot weight and root growth) as compared with 50 g L^-1 PEG selected somaclone lines under limited irrigation. The yield parameters-panicle length, panicle weight, grains per panicle, 1,000-grain weight, grain yield per plant, harvest index and grain straw ratio were also higher in 30 g L^-1 PEG lines as compared with 50 g L^-1 PEG lines. The results, therefore indicate that 30 g L^-1 PEG selected somaclone lines were more suited than 50 g L^-1 PEG selected somaclone lines under stress as compared with WT. The finding suggests that rice cv PR113 somaclones generated on PEG are found to be drought tolerant under field condition with better yield.     

In vitro strategies for selection of eye-spot resistant sugarcane lines using toxins of Helminthosporium sacchari

In vitro strategies were applied for the selection of eye-spot resistant variants from susceptible sugarcane cultivar Co 419 Different selective units (callus and leaf) of the susceptible cultivar were subjected to sub-lethal to lethal doses of toxins (culture filtrate and partially purified toxin) of H. sacchari, with the objective of improving the efficacy of in vitro selection protocols. All the selective units gave more or less similar response with culture filtrate, but a distinct response was observed when leaf was subjected to partially purified toxin treatment. The response was characterised by the degree of resistance exhibited by the regenerated seedlings.     

In vitro leaf-derived organogenesis and somaclonal variant detection in Humulus lupulus L

In Vitro Cellular & Developmental Biology - Plant - The exploitation of somaclonal variation potentially could be a valid strategy to overcome the depletion of hop intraspecific...     

In vitro generation of an HTLV-III variant by neutralizing antibody

Transmission and culture of "parental" virus (HTLV-III) from H9 cells transfected with the cloned isolate (lambda HXB-2D) in human serum possessing HTLV-III neutralizing antibody selected for a "variant" that was not neutralized by the selecting serum but was neutralized by another antibody-positive serum "Control" virus, selected in serum lacking neutralizing antibody, and the variant showed highly similar tryptic peptide maps of the major envelope glycoprotein, and no changes in restriction enzyme patterns of viral DNA. These findings show that HTLV-III type-specific neutralizing antibodies occur, can influence the propagation of variant viruses that may arise, and presumably result from minor changes in the eliciting antigen. The extent to which such type-specific neutralizing antibodies influence immune surveillance against HTLV-III infection in vivo, a question with relevance to future vaccination attempts, remains to be determined. Nucleotide sequencing of the control and variant envelope genes may elucidate a region important for virus neutralization and vaccine development.     

Description of Gibberella sacchari and neotypification of its anamorph Fusarium sacchari

We described the teleomorph of Fusarium sacchari as Gibberella sacchari, sp. nov. This species can be separated from other species of Gibberella on the basis of the longer, narrower ascospores found in G. sacchari and by sexual cross fertility. Female-fertile mating type tester strains were developed that can be used for making sexual crosses with this heterothallic fungus under laboratory conditions. The anamorph, Fusarium sacchari, was neotypified.     

Resistance to Melanaphis sacchari in the sugarcane cultivar R 365

This study focuses on the resistance of sugarcane, Saccharum spec. (Poaceae), to the sugarcane aphid, Melanaphis sacchari (Zehntner) (Hemiptera: Aphididae), which vectors Sugarcane yellow leaf virus (SCYLV). Resistance was characterized in cultivar R 365, using a 3‐year field trial and laboratory experiments on potted plantlets and excised leaves. R 365 reduced aphid populations in the field by antixenosis and antibiosis. Using the electrical penetration graph technique, we detected delayed aphid salivation in phloem and inhibition of passive phloem sap uptake in R 365. The resistance factors also proved to be effective against the corn leaf aphid, Rhopalosiphum maidis (Fitch) (Hemiptera: Aphididae), another vector of SCYLV.     

Identification and genetic diversity of Fusarium sacchari associated with pokkah boeng disease of sugarcane in Northeast Brazil

Isolates from the Fusarium fujikuroi species complex, mainly F. sacchari, have been reported to be the causal agents of pokkah boeng in sugarcane in Brazil. However, inadequate information was available on the occurrence and genetic diversity of F. sacchari in Northeast Brazil, which is a limiting factor on management. Thus, isolates of F. subglutinans sensu lato from sugarcane plants with symptoms of pokkah boeng were evaluated using the sexual cross-fertility to determine species. All the isolates produced black perithecia when they were crossed with the test isolates of F. sacchari. Three weeks after the crossing, the formation of fertile ascospores cirri was observed. Thirty-four isolates were self-sterile hermaphrodites, while 21 were fertile only as males. Five isolates were homothallic. The effective size [N_e(f)] of the population as a function of the frequency of hermaphrodites and female sterile strains was 95.5%. The F. sacchari isolates were separated into four genetic groups independent of geographic location. The mean of polymorphism among all populations was 79%, and the average unbiased genetic diversity (uh) was considered moderate (0.31). This study in addition to confirming that F. sacchari as the main species associated with pokkah boeng in sugarcane in Northeast Brazil, reveals the relationship of mating type and genetic diversity of F. sacchari. The unrestricted gene flow between regions is probably the best explanation for the low geographic correlation. This knowledge will help in the adoption of management measures with fungicides or resistant cultivars.     

In vitro selection of alfalfa plants resistant to Fusarium oxysporum f. sp. medicaginis

Summary From two lines of Medicago sativa characterized by a high regeneration capability, calli resistant to culture filtrate of Fusarium oxysporum f. sp. medicaginis have been selected. In these calli regeneration capability was greatly reduced and only one plant per callus was recovered. Regenerated plants have been evaluated for resistance to culture filtrate and for in vivo resistance to the pathogen. Three plants out of eight were resistant to the fungus and a high correlation between resistance to culture filtrate and in vivo resistance was observed.Research work supported by C.N.R., Italy. Special grant I.P.R.A. Subproject 1, paper no. 1468     

In vitro culture and the incidence of somaclonal variation in regenerated plants of Trifolium pratense L.

Red clover (Trifolium pratense L.) cvs ‘Altaswede’ (2n = 2x = 14) and ‘Norseman’ (2n = 4x = 28) have been used to investigate tissue culture initiation, plant regeneration and the occurrence of somaclonal variation. After callus induction shoots were induced both when calli on L2 medium containing 2 mg l⁻¹ 2,4-dichlorophenoxy acetic acid (2,4-D), 2 mg l⁻¹ 6-benzylaminopurine (BA) and 2 mg l⁻¹ 6-amino-purine (AP) were subcultured on media containing naphthalene acetic acid (NAA) (0.05 mg l⁻¹) and kinetin (KIN) (0.05 or 0.5 mg l⁻¹) and when embryogenic calli were cultured and subcultured on L2 medium containing 0.002 mg l⁻¹ 4-amino-3,5,6-trichloropicolinic acid (PIC) and 0.2 mg l⁻¹ BA. Shoot tip cultures were also established to induce multiple shoots for regeneration of plants via organogenesis.Regenerants from different regeneration pathways were evaluated for chromosome number stability, morphology and several biochemical traits. Regenerated plants showed stable isozyme banding patterns for malate dehydrogenase, 6-phosphogluconate dehydrogenase, phosphoglucose isomerase, phosphoglucomutase and shikimate dehydrogenase, as well as their nodule leghemoglobin profiles. Variations were detected in the chromosome number of some regenerants as well as in leaflet length-to-width ratio and leaflet number. Factors related to the incidence of somaclonal variation are discussed.     

In vitro selection for salt tolerance in crop plants: Theoretical and practical considerations

Summary In recent years attempts have been made to supplement traditional breeding for the production of salt-tolerant plants with variability existing in cell culture. The potential causes suggested as an explanation for the limited success of the in vitro approach include: a) lack, or loss during selection, of regeneration capability; b) the development of epigenetically adapted cells; c) lack of correlation between the mechanisms of tolerance operating in cultured cells and mechanisms that operate in cells in the intact plant; and d) multigenicity of salt tolerance. The recent successful production of healthy, fertile, and genetically stable salt-tolerant regenerants from cells obtained from highly morphogenic explants which are selected early in culture (using one-step or short-term strategies) for salt tolerance, together with the demonstration that salt-sensitive plants can become tolerant by mutations in one or few genes, suggest that some of the potential limitations can be overcome and that some of them may not exist at all.     

Binomial sequential sampling of adult Saccharicoccus sacchari on sugarcane

The between-stalk dispersion characteristics of adults of the pink sugarcane mealybug Saccharicoccus sacchari (Cockerell) were determined in southern Queensland. Iwao's patchiness regression was inappropriate to describe the relationship between mean and variance. Taylor's power law indicated that adults were aggregated, especially at the beginning and end of the ratoon growth period. Binomial data were modelled by the Nachman model; the model of Grout and two models of Wilson & Room were inappropriate to describe the relationship between proportion of stalks infested and mean numbers of adults per stalk. Relationships to determine sample sizes for fixed levels of precision and binomial fixed-precision-level stop lines are developed for different sampling times using Taylor's power law and Nachman's equation.     

In vitro biopesticide effect of alkaloids and flavonoids of some plants against Fusarium oxysporum

Alkaloids and flavonoids of three plants, namely Withania somnifera, Euphorbia hirta and Terminalia chebula, were tested for their antifungal potential against plant pathogenic fungi Fusarium oxysporum. The spore counting technique was followed for the antibiotic properties of the extracts tested at three different concentrations (2.5, 5 and 10?mg/ml). Results revealed that the bound flavonoid of root of W. somnifera and free flavonoids of stem bark and fruits of T. chebula totally inhibited spore germination of the fungi (100%). The range of minimum inhibitory concentration and minimum fungicidal concentration of extracts recorded was 0.039–0.625 and 0.039–1.25?mg/ml, respectively. Bound flavonoids of selected plants were more effective than free flavonoids and alkaloids. The study suggests that both alkaloids and flavonoids extracts of selected plants have a potential antifungal properties for the control of diseases of various crops.     

In vitro saprotrophic basidiomycetes tolerance to pendimethalin

Pendimethalin is a dinitroaniline herbicide classified among the persistent-bioaccumulative toxics. In this paper, the tolerance to this herbicide has been studied in isolates of basidiomycetes (10 species including 9 wood-rotting and 1 litter fungi), collected in different areas of the Campania region (South Italy). The isolates were grown on two different agar media, rich and poor for the presence/absence of dextrose and NH₄NO₃, and amended with 0, 100 and 500 ppm herbicide. The mycelial growth was recorded daily, and statistical analysis of fungal growth rates, determined via linear regression, allowed us to compare the hyphal extension of various macrofungi in presence of herbicide. These data represent the adaptation capacity of the fungal organisms to the peculiar environmental situation. In amended agar, all fungi exhibited a certain tolerance to pendimethalin and, normally, the fungal growth decreased with the increasing of pollutant concentration. Nevertheless, in some cases, the difference of ground reflected the different agar media. In fact, the growth of Agrocybe aegerita, in rich agar decreased with the increase of herbicide dose, while in poor agar it increased with the pollutant. Among the examined fungi, A. aegerita was the species which resulted the most tolerant to herbicide, showing growth to about 70% of the control to the highest concentrations of pollutant and for the two different agar media.Scientific relevance of the paperNo published report is available regarding pendimethalin herbicide biodegradation with basidiomycetes; therefore this work represented the first and preliminary investigation as regards. It's directed to determine the fungal tolerance to the pollutant.     

Identification of somaclonal variants of sugarcane (Saccharum spp.) resistant to sugarcane mosaic virus via RAPD markers

Somaclonal variants resistant to sugarcane mosaic virus (SCMV) were obtained from susceptible sugarcane cv PR62258 through somatic embryogenesis by increasing the number of subcultures of the embryogenic callus tissue in MS medium with 3 mg/L 2,4-dichlorophenoxyacetic acid. Transfers were made at 30-day intervals for 1, 2 or 3 subcultures. Two somaclones, namely AT626 and BT627, were selected by their resistance to SCMV. These subclones have maintained the resistance trait over seven years of testing in the field. In this report we identified the somaclonal SCMV resistant variants from the maternal line and the nonresistant somaclones, using the RAPD technique.     

Analysis of aMusa spp. somaclonal variant resistant to yellow Sigatoka

Somaclonal variant CIEN BTA-03 resistant to yellow Sigatoka was obtained from a susceptible banana clone (Williams clone), by increasing the production of adventitious buds using 6-Belcilaminopurine at high concentrations. This somaclone has exhibited yellow Sigatoka resistance in the field for five consecutive years of asexual reproduction. We used RAPD markers to generate characteristic “fingerprints” for each probe, concluding that they are reliable tools for evaluating the genetic variability ofMusa regenerants obtained byin vitro culture.     

In vitro selection for Fusarium wilt resistance in Gladiolus

Cormels pieces of four Fusarium susceptible Gladiolus cultivara (Friendship, Peter Pears, Victor Borge and Novalux) formed friable calli when cultured in vitro on Murashige and Skoog basal medium containing various concentrations of auxin and cytokinin. The friable calli established cell suspensions. Plantlet regeneration was obtained from the control callus, control cell suspension derived callus and in vitro selected Fusarium oxysporum Schlecht. resistant cell-lines of Friendship. The in vitro cormlets showed 85-95% germination after breaking dormancy of 8weeks at 4℃. Cell suspensions of all four Gladiolus cultivara were found to be highly sensitive to fusaric acid. Gradual Increase in fusaric acid concentrations to the cell-suspension cultures decreased cell growth considerably. One albino plant was found from the second generation of the In vitro selected cell line of Friendship. The albino plant was found to be highly susceptible to F. oxysporum. The cormlets of all in vitro selected call lines of Friendship were inoculated with a conidial suspension of the F. oxysporum before planting and were also sprayed with the same spore suspension for further characterization when the height of plants was about 6cm. The four selected cell lines showed the same response whether or not they were Inoculated with conidia of the F. oxysporum. Plantlets of all of the selected call lines exhibited significant growth as compared with the control after application of conidia of the F.oxysporum.     

Epigenetic aspects of somaclonal variation in plants

Somaclonal variation is manifested as cytological abnormalities, frequent qualitative and quantitative phenotypic mutation, sequence change, and gene activation and silencing. Activation of quiescent transposable elements and retrotransposons indicate that epigenetic changes occur through the culture process. Epigenetic activation of DNA elements further suggests that epigenetic changes may also be involved in cytogenetic instability through modification of heterochromatin, and as a basis of phenotypic variation through the modulation of gene function. The observation that DNA methylation patterns are highly variable among regenerated plants and their progeny provides evidence that DNA modifications are less stable in culture than in seed-grown plants. Future research will determine the relative importance of epigenetic versus sequence or chromosome variation in conditioning somaclonal variation in plants.     

Investigation into the ability of Gluconacetobacter sacchari to live as an endophyte in sugarcane

The relatively low numbers and sporadic pattern of incidence of the acetic acid bacterium Gluconacetobacter sacchari with the pink sugarcane mealybug (PSMB) Saccharicoccus sacchariCockerell (Homoptera: Pseudococcidae) over time and from different sugarcane-growing regions do not indicate that Glac. sacchari is a significant commensal of the PSMB, as has been previously proposed. This study was conducted to investigate the hypothesis that Glac. sacchari is, like its closest relative Glac. diazotrophicus, an endophyte of sugarcane (Saccharum officinarum L.). In this study, bothGlac. sacchari and Glac. diazotrophicus were isolated from internal sugarcane tissue, although the detection of both species was sporadic in all sugarcane-growing regions of Queensland tested. To confirm the ability of Glac. sacchari to live endophytically, an experiment was conducted in which the roots of micropropagated sugarcane plantlets were inoculated with Glac. sacchari, and the plantlets were subsequently examined for the presence of the bacterium in the stem cells. Pure cultures of Glac. sacchari were grown from homogenized surface sterilized sugarcane stems inoculated withGlac. sacchari.Electron microscopy was used to provide further conclusive evidence that Glac. sacchari lives as an endophyte in sugarcane. Scanning electron microscopy of (SEM) sugarcane plantlet stems revealed rod-shaped cells of Glac. sacchari within a transverse section of the plantlet stem cells. The numbers of bacterial cells inside the plant cell indicated a successful infection and colonization of the plant tissue. Using transmission electron microscopy, (TEM) bacterial cells were more difficult to find, due to their spatial separation. In our study, bacteria were mostly found singularly, or in groups of up to four cells inside intercellular spaces, although bacterial cells were occasionally found inside other cells.