In Vitro Anticancer Activity of Staphyloxanthin Pigment Extracted from <Emphasis Type="Italic">Staphylococcus gallinarum</Emphasis> KX912244, a Gut Microbe of <Emphasis Type="Italic">Bombyx mori</Emphasis>

The present study reports the in vitro biological nature of the pigment produced by Staphylococcus gallinarum KX912244, isolated as the gut microflora bacterium of the insect Bombyx mori. The purified pigment was characterized as Staphyloxanthin based on bio-physical characterization techniques like Fourier transform infrared spectroscopy, high performance liquid chromatography, Proton nuclear magnetic resonance spectroscopy (¹H NMR), Liquid chromatography-Mass spectroscopy and Gas chromatography-Mass spectroscopy. The Staphyloxanthin pigment presented considerable biological properties including in vitro antimicrobial activity against pathogens Staphylococcus aureus, Escherichia coli and Candida albicans; in vitro antioxidant activity by % DPPH free radical scavenging activity showing IC₅₀ value of 54.22 µg/mL; DNA damage protection activity against reactive oxygen species and anticancer activity evaluated by cytotoxicity assay against 4 different cancer cell lines like the Dalton’s lymphoma ascites with IC₅₀ value 6.20?±?0.02 µg/mL, Ehrlich ascites carcinoma having IC₅₀ value 6.48?±?0.15 µg/mL, Adenocarcinomic human alveolar basal epithelial cells (A549 Lung carcinoma) bearing IC₅₀ value 7.23?±?0.11 µg/mL and Mus mucus skin melanoma (B16F10) showing IC₅₀ value 6.58?±?0.38 µg/mL and less cytotoxicity towards non-cancerous human fibroblast cell lines (NIH3T3) with IC₅₀ value of 52.24 µg/mL. The present study results suggest that Staphyloxanthin acts as a potential therapeutic agent especially due to its anticancer property.     

Novel rhodanine derivatives are selective algicides against <Emphasis Type="Italic">Microcystis aeruginosa</Emphasis>

In this study, a series of rhodanine derivatives containing various substituents was synthesized and tested for in vitro algicidal activity. Among the tested substituent groups, phenyl substituents with halogen groups showed good inhibitory potency. Furthermore, the compound with chlorine at the C2 position of the phenyl ring exhibited a higher algicidal effect than the compound with chlorine at the C3 position of the phenyl ring. Among the various rhodanine derivatives tested, 5-(2,4-dichlorobenzylidene)- rhodanine (compound 20) was the most potent inhibitor against M. aeruginosa with a lethal concentration 50 (LC₅₀) value of 0.65 μM and Selenastrum capricornutum with an LC₅₀ value of 0.82 μM. To verify the feasibility of their use in ecosystems, 25 h of acute ecotoxicity tests were carried out for three derivatives against Danio rerio and Daphnia magna. No mortality was observed in groups exposed to 2.0 μM of compound 20 after 100 h of exposure. Moreover, a survival rate of 100% was observed in D. magna exposed to 2 μM of compound 20 for 100 h. Overall, the results show that rhodanine derivatives are a possible method for controlling and inhibiting harmful algal blooms.     

Antibiofilm potential of biogenic silver nanoparticles against <Emphasis Type="Italic">Kocuria rosea</Emphasis> And <Emphasis Type="Italic">Kocuria rhizophila</Emphasis>

The present study aims at biosynthesizing, characterizing and evaluating the biogenic silver nanoparticles (AgNPs) as antimicrobial and antibiofilm against Kocuria rosea and Kocuria rhizophila. Cellfree supernatant of Proteus mirabilis culture was used for biosynthesizing AgNPs, which confirmed by visualizing color change and X-ray diffraction. Transmission electron microscopy showed the formation of AgNPs in the range of 5–40 nm. ART-FTIR spectra provided evidence for presence of proteins as possible biomolecules responsible for stability of AgNPs and act as capping agent. AgNPs had ability to inhibit growth of K. rosea and K. rhizophila. The minimum inhibitory concentration (MIC₉₀) of AgNPs against both strains was 25 μg/mL. Antiadhesive effect of AgNPs was verified at sub-MIC₉₀ dose (12.5 μg/mL). The AgNPs concentrations up to 100 μg/mL were not effective for complete removing the already established biofilms with maximum removing percentage of 30.5–34.9%. In conclusion, the present study demonstrated an unprecedented green process for biosynthesizing stable spherical-shaped AgNPs. Early control is suggested by preventing biofilm formation using low AgNPs concentration (12.5 μg/mL) as a potential ingredient for formulating effective chemical sanitizers.     

Nematicidal metabolites from <Emphasis Type="Italic">Gliocladium roseum</Emphasis> YMF1.00133

A strain of the fungus Gliocladium roseum YMF1.00133 was found to secrete nematicidal metabolites against nematodes Panagrellus redivivus, Caenothabditis elegans and Bursaphelenchus xylophilus in experiments searching for nematicidal fungi. Through bioassay-guided fractionations, a unique trioxopiperazine alkaloid, gliocladin C (compound 1), and an alkylane resorcinol, 5-n-heneicosylresorcinol (compound 2) were obtained from the methanol extract of the fungus and determined by single-crystal X-ray analysis and spectroscopic data. In vitro immersion experiments showed that the ED₅₀ values of compounds 1 and 2 after 24 h incubation were 15 and 30 μg/mL against C. elegans, 50 and 80 μg/mL against P. redivivus, and 200 and 180 μg/mL against B. xylophilus, respectively. The X-ray diffraction data of compound 1 and the nematicidal activity of compounds 1 and 2 were reported for the first time.     

Expression of a rice <Emphasis Type="Italic">GLP</Emphasis> in <Emphasis Type="Italic">Medicago truncatula</Emphasis> exerting pleiotropic effects on resistance against <Emphasis Type="Italic">Fusarium oxysporum</Emphasis> through enhancing FeSOD-like activity

To evaluate the effectiveness of a germin-like protein (GLP) in legumes against the serious soil-borne pathogen Fusarium oxysporum f. sp. lentis, an Oryza sativa root-expressed GLP (OsRGLP1) was expressed in the model legume Medicago truncatula using the recombinant vector pCOsRGLP1. The transgene was highly expressed in M. truncatula transformed lines as assessed by RT-qPCR. Consistent with the active status of the transgene there was an elevated accumulation of H₂O₂ in transformed progeny. Enzymatic characterization of T₁ transgenic progeny showed increased superoxide dismutase (SOD) activity. The additional SOD activity in transgenic lines was insensitive to potassium cyanide and sensitive to H₂O₂ indicating its resemblance to FeSOD. The effectiveness of the OsRGLP1 gene was tested by monitoring the root disease after infection of wild-type and transgenic lines. Wild-type plants were greatly affected by the pathogen infection showing a percent disease index value of 50 compared to 10–18 for the transgenic lines. The tolerance of the transgenic lines leads to recovery in fresh weight and pod production to an almost normal level. Analysis of defense-related genes downstream of hydrogen peroxide (H₂O₂) in transgenic plants showed induction of salicylic acid and jasmonate signaling pathways and increased expression of some pathogenesis-related-1 (PR-1) genes and a plant defensin gene. Overall, the findings suggest that OsRGLP1 provides protection against the fungal pathogen F. oxysporum that may involve the direct influence of H₂O₂ on signaling pathways leading to the activation of defense-related genes.     

<Emphasis Type="Italic">Nicotiana sylvestris</Emphasis> L. mutants resistant to isopropyl <Emphasis Type="Italic">N</Emphasis>-phenyl carbamate possess microtubule organizing centers of heightened stability

N. sylvestris mutants resistant to isopropyl N-phenyl carbamate (IPC), a herbicide belonging to the phenyl carbamate series, are obtained by means of in vitro selection using gamma radiation. A concentration of 30 μM IPC was found to be the maximum concentration at which mutants of the N. sylvestris line capable of regeneration and rooting under conditions of selection pressure could be selected. IPC resistance in the mutants obtained was confirmed by a number of tests, in particular, tests that measure the capacity of leaf explants of the mutant lines to regenerate plants and the ability of their callus cells to survive in media with a selective IPC concentration, as well as by means of genetic, morphometric, cytological, and immunofluorescent analyses. The results of these studies attest to increased resistance of the mutant plants to this antimitotic substance by comparison with a control. It is shown that resistance to IPC is based on the heightened resistance of the microtubule organizing centers of the cells of these lines. It is established that the acquired resistance trait inherited in the F₁ and F₂ generations of the mutants is a dominant nuclear trait.     

Over-expressing a UDP-glucosyltransferase gene (<Emphasis Type="Italic">Ta-UGT</Emphasis><Subscript><Emphasis Type="Italic">3</Emphasis></Subscript>) enhances <Emphasis Type="Italic">Fusarium</Emphasis> Head Blight resistance of wheat

Wheat Fusarium Head Blight (FHB), mainly caused by Fusarium graminearum (F.g), is a destructive fungal disease worldwide. FHB can not only cause considerable reduction in yield, but more seriously, can contaminate grain by trichothecene toxins released by the fungus. Here, we report new insights into the function and underlying mechanisms of a UDP-glycosyltransferase gene, Ta-UGT ₃ , that is involved in FHB resistance in wheat. In our previous study, Ta-UGT ₃ was found to enhance host tolerance against deoxynivalenol (DON) in Arabidopsis. In this study, four transgenic lines over-expressing Ta-UGT ₃ in a FHB highly susceptible wheat variety, Alondra’s, were obtained and characterized. 3 years of assays using single floret inoculation with F.g indicated that all four transgenic lines exhibited significantly enhanced type II resistance to FHB and less DON accumulation in the grains compared to the untransformed control. Histological observation using GFP labelled F.g was in agreement with the above test results since over-expression of Ta-UGT ₃ dramatically inhibited expansion of F.g. To explore the putative mechanism of resistance mediated by Ta-UGT ₃ , microarray analysis, qRT-PCR and hormone measurements were performed. Microarray analysis showed that DON up-regulated genes, such as TaNPR1, in the susceptible control, and down-regulated genes in F.g inoculated transgenic lines, while qRT-PCR showed that some defence related genes were up-regulated in F.g inoculated transgenic lines. Ta-UGT ₃ over-expression also changed the contents of the endogenous hormones SA and JA in the spikes. These data suggest that Ta-UGT ₃ positively regulates the defence responses to F.g, perhaps by regulating defence-related and DON-induced downstream genes.     

Overexpression of <Emphasis Type="Italic">MuHSP70</Emphasis> gene from <Emphasis Type="Italic">Macrotyloma uniflorum</Emphasis> confers multiple abiotic stress tolerance in transgenic <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis>

A 70-KD heat shock protein (HSP70) is one of the most conserved chaperones. It is involved in de novo protein folding and prevents the aggregation of unfolded proteins under lethal environmental factors. The purpose of this study is to characterise a MuHSP70 from horsegram (Macrotyloma uniflorum) and elucidating its role in stress tolerance of plants. A MuHSP70 was cloned and characterised from a natural drought stress tolerant HPK4 variety of horsegram (M. uniflorum). For functional characterization, MuHSP70 was overexpressed in transgenic Arabidopsis. Overexpression of MuHSP70 was found to provide tolerance to the transgenic Arabidopsis against various stresses such as heat, cold, drought, salinity and oxidative stress. MuHSP70 transgenics were observed to maintain the shoot biomass, root length, relative water content, and chlorophyll content during exposure to multi-stresses relative to non-transgenic control. Transgenic lines have further shown the reduced levels of MDA, H₂O₂, and proteolytic activity. Together, these findings suggest that overexpression of MuHSP70 plays an important role in improving abiotic stress tolerance and could be a crucial candidate gene for exploration in crop improvement program.     

Dormant forms of <Emphasis Type="Italic">Micrococcus luteus</Emphasis> and <Emphasis Type="Italic">Arthrobacter globiformis</Emphasis> not platable on standard media

The colony-forming ability of long (3–9 months) incubated cystlike resting cells (CRC) of the nonspore-forming gram-positive bacteria Micrococcus luteus and Arthrobacter globiformis was studied in this work. The preservation of the CRC proliferative potential as assayed by plating on standard LB agar was shown to depend on the conditions of the formation of the dormant cells. In aged post-stationary cultures of micrococci and arthrobacters grown under carbon and phosphorus limitation the number of colony-forming units (CFU/ml) of CRC decreased in the course of 3–9 month incubation to the level of 10⁶–10⁷ CFU/ml. However, M. luteus CRC obtained under carbon and nitrogen limitation and A. globiformis CRC obtained under nitrogen limitation and starvation completely lost their ability to form colonies on standard solid medium after 4–6 months of incubation and turned into a ‘non-culturable’ (non-platable) state. In this case, the ratio of live cells in the population of M. luteus and A. globiformis ‘non-culturable’ CRCs (determined by the Live/Dead staining test) was 10–44% of the total cell number. To study the possible preservation of proliferative potential in non-platable CRCs, various methods of their reactivation were applied. Although preincubation of CRC suspensions in a buffer solution of 0.1 M K₂HPO₄ (pH 7.4) or in the presence of lysozyme (1 or 10 μg/ml) resulted in increased numbers of live cells (determined by the Live/Dead test) or in disruption of the cell conglomerates, it did not increase considerably the CFU titer on LB medium. Variations in the medium composition, such as addition of sodium pyruvate as an antioxidant or dilution of the medium, promoted the formation of macrocolonies by a small portion of nonplateable CRC of M. luteus (50?80 CFU/ml), whereas the number of the cells capable of microcolony formation (mCFU) was 1.8–6.8 × 10⁵ mCFU/ml, exceeding the CFU titers by four orders of magnitude. The application of semisolid agar and the most probable number (MPN) method was the most efficient for determination of the mCFU titer, and an almost complete reversion of ‘non-culturable’ micrococcal CRCs to microcolony formation was observed (up to 2.3 × 10⁷ mCFU/ml). The usefulness of diluted complete media for the restoration of the colony-forming ability of the dormant forms was confirmed in experiments with ‘nonculturable’ CRCs of A. globiformis. The development of special procedures and methods for determining actively proliferating cells not detected by ordinary methods is of great importance for advanced monitoring studies.     

<Emphasis Type="Italic">STAYGREEN</Emphasis> (<Emphasis Type="Italic">CsSGR</Emphasis>) is a candidate for the anthracnose (<Emphasis Type="Italic">Colletotrichum orbiculare</Emphasis>) resistance locus <Emphasis Type="Italic">cla</Emphasis> in Gy14 cucumber

Key message

Map-based cloning identified a candidate gene for resistance to the anthracnose fungal pathogen Colletotrichum orbiculare in cucumber, which reveals a novel function for the highly conserved STAYGREEN family genes for host disease resistance in plants.

Abstract

Colletotrichum orbiculare is a hemibiotrophic fungal pathogen that causes anthracnose disease in cucumber and other cucurbit crops. No host resistance genes against the anthracnose pathogens have been cloned in crop plants. Here, we reported fine mapping and cloning of a resistance gene to the race 1 anthracnose pathogen in cucumber inbred lines Gy14 and WI 2757. Phenotypic and QTL analysis in multiple populations revealed that a single recessive gene, cla, was underlying anthracnose resistance in both lines, but WI2757 carried an additional minor-effect QTL. Fine mapping using 150 Gy14?×?9930 recombinant inbred lines and 1043 F₂ individuals delimited the cla locus into a 32 kb region in cucumber Chromosome 5 with three predicted genes. Multiple lines of evidence suggested that the cucumber STAYGREEN (CsSGR) gene is a candidate for the anthracnose resistance locus. A single nucleotide mutation in the third exon of CsSGR resulted in the substitution of Glutamine in 9930 to Arginine in Gy14 in CsSGR protein which seems responsible for the differential anthracnose inoculation responses between Gy14 and 9930. Quantitative real-time PCR analysis indicated that CsSGR was significantly upregulated upon anthracnose pathogen inoculation in the susceptible 9930, while its expression was much lower in the resistant Gy14. Investigation of allelic diversities in natural cucumber populations revealed that the resistance allele in almost all improved cultivars or breeding lines of the U.S. origin was derived from PI 197087. This work reveals an unknown function for the highly conserved STAYGREEN (SGR) family genes for host disease resistance in plants.

     

In vitro anticancer properties of selected <Emphasis Type="Italic">Eucalyptus</Emphasis> species

In spite of the recent advancements in oncology, the overall survival rate for pancreatic cancer has not improved over the last five decades. Eucalypts have been linked with cytotoxic and anticancer properties in various studies; however, there is very little scientific evidence that supports the direct role of eucalypts in the treatment of pancreatic cancer. This study assessed the anticancer properties of aqueous and ethanolic extracts of four Eucalyptus species using an MTT assay. The most promising extracts were further evaluated using a CCK-8 assay. Apoptotic studies were performed using a caspase 3/7 assay in MIA PaCa-2 cells. The aqueous extract of Eucalyptus microcorys leaf and the ethanolic extract of Eucalyptus microcorys fruit inhibited the growth of glioblastoma, neuroblastoma, lung and pancreatic cancer cells by more than 80% at 100 μg/mL. The E. microcorys and Eucalyptus saligna extracts showed lower GI₅₀ values than the ethanolic Eucalyptus robusta extract in MIA PaCa-2 cells. Aqueous E. microcorys leaf and fruit extracts at 100 μg/mL exerted significantly higher cell growth inhibition in MIA PaCa-2 cells than other extracts (p < 0.05). Statistically similar IC₅₀ values (p > 0.05) were observed in aqueous E. microcorys leaf (86.05 ± 4.75 μg/mL) and fruit (64.66 ± 15.97 μg/mL) and ethanolic E. microcorys leaf (79.30 ± 29.45 μg/mL) extracts in MIA PaCa-2 cells using the CCK-8 assay. Caspase 3/7-mediated apoptosis and morphological changes of cells were also witnessed in MIA PaCa-2 cells after 24 h of treatment with the extracts. This study highlighted the significance of E. microcorys as an important source of phytochemicals with efficacy against pancreatic cancer cells. Further studies are warranted to purify and structurally identify individual compounds and elucidate their mechanisms of action for the development of more potent and specific chemotherapeutic agents for pancreatic cancer.     

Comparative study between Larval Packet Test and Larval Immersion Test to assess the effect of <Emphasis Type="Italic">Metarhizium anisopliae</Emphasis> on <Emphasis Type="Italic">Rhipicephalus microplus</Emphasis> tick larvae

Entomopathogenic fungi, such as Metarhizium anisopliae, for the control of arthropods, have been studied for more than 20 years. The aim of this study was to determine the best methodology to evaluate the in vitro effect of the fungus M. anisopliae on Rhipicephalus microplus tick larvae. We compared a modified Larval Packet Test (LPT) and a Larval Immersion Test (LIT). For the LPT filter papers were impregnated with 1 mL of M. anisopliae suspension in Triton X-100 at 0.02%, in concentrations of 10⁶, 10⁷ and 10⁸ conidia/mL and subsequently folded to include the larval ticks. LIT was performed by immersing the larvae in M. anisopliae suspensions for 5 min using the same three concentrations, then the larvae were placed on filter paper clips. For LPT, the LT₅₀ values obtained were 134.6, 27.2 and 24.8 days for concentrations of 10⁶, 10⁷ and 10⁸ conidia/mL; and the mortality after 21 days was 17.3, 17.6 and 38%, respectively. The LT₅₀ values of LIT were 24.5, 20 and 9.2 days with mortality after 21 days of 50.5, 64.7 and 98% for 10⁶, 10⁷ and 10⁸ conidia/mL, respectively. For the same conidia concentration, LIT showed a higher mortality in a shorter time interval when compared with LPT. These differences between the methods tested must be taking into account in further screening and effect studies with M. anisopliae. The set of results shown here could optimize the protocol used to identify M. anisopliae strains pathogenic against R. microplus.     

Aluminium pre-treatment induces activation of defense responses against <Emphasis Type="Italic">Fusarium</Emphasis> infection in <Emphasis Type="Italic">Triticum aestivum</Emphasis>

Pre-treatment with low dose of aluminium sulphate [Al₂(SO₄)₃] was assayed on wheat (Triticum aestivum L.) seedlings for its ability to induce resistance against Fusarium oxysporum infection. Pre-treatment of seven days old germinated wheat seedlings with 50 μM concentration of Al₂(SO₄)₃ reduced the severity of the disease. In Al₂(SO₄)₃ pre-treated seedlings inoculated with fungus reduction in disease severity was correlated with suppression of fungal mycelia development inside the leaf tissues. The activities of peroxidase, polyphenol oxidase, and phenylalanine ammonia lyase were determined as resistance markers. F. oxysporum inoculation induced significant increase of all these enzymes. Such responses were expressed earlier and with much higher magnitude when Al₂(SO₄)₃ pre-treated seedlings were challenged with the pathogen. Slower disease development in Al₂(SO₄)₃ pre-treated seedlings might be due to increased deposition of total phenolic compounds and enhanced level of salicylic acid which restricts pathogen entry.     

The <Emphasis Type="Italic">in vitro</Emphasis> and <Emphasis Type="Italic">in vivo</Emphasis> efficacy of fluconazole in combination with farnesol against <Emphasis Type="Italic">Candida albicans</Emphasis> isolates using a murine vulvovaginitis model

Farnesol is a quorum-sensing molecule that inhibits biofilm formation in Candida albicans. Previous in vitro data suggest that, in combination with certain antifungals, farnesol may have an adjuvant anti-biofilm agent. However, the in vivo efficacy of farnesol is very questionable. Therefore, the in vitro and in vivo activity of fluconazole combined with farnesol was evaluated against C. albicans biofilms using fractional inhibitory concentration index (FICI) determination, time-kill experiments and a murine vulvovaginitis model. The median biofilm MICs of fluconazole-sensitive C. albicans isolates ranged between 4 -> 512 mg/L and 150–300 μM for fluconazole and farnesol, respectively. These values were 512 -> 512 mg/L and > 300 μM for fluconazole-resistant clinical isolates. Farnesol decreased the median MICs of fluconazole by 2-64-fold for biofilms. Based on FICI, synergistic interaction was observed only in the case of the sessile SC5314 reference strain (FICIs: 0.16–0.27). In time-kill studies, only the 512 mg/L fluconazole and 512 mg/L fluconazole + 75 μM farnesol reduced biofilm mass significantly at each time point in the case of all isolates. The combination reduced the metabolic activity of biofilms for all isolates in a concentration- and time-dependent manner. Our findings revealed that farnesol alone was not protective in a murine vulvovaginitis model. Farnesol was not beneficial in combination with fluconazole for fluconazole-susceptible isolates, but partially increased fluconazole activity against one fluconazole-resistant isolate, but not the other one.     

<Emphasis Type="Italic">Diplazium</Emphasis> hybrids involving <Emphasis Type="Italic">D. plantaginifolium</Emphasis> and <Emphasis Type="Italic">D</Emphasis>. <Emphasis Type="Italic">ternatum</Emphasis> from Mexico and Central America

Here we evaluate the origins and relationships of Mexican and Central American Diplazium hybrids derived from crosses involving either D. plantaginifolium or D. ternatum. Based on study of live plants and herbarium specimens, we distinguish D. ×verapax from the similar D. riedelianum and present evidence that the former is a sterile hybrid derived from a cross between D. plantaginifolium and D. werckleanum. We also describe new hybrids, D. ×torresianum and D. ×subternatum from Mexico and northern Central America. Both involve D. ternatum as one parent. Diplazium. cristatum is the other putative parent of D. ×torresianum, and D. plantaginifolium is the second parent of D. ×subternatum. We also designate lectotypes for D. cordovense and D. dissimile.     

Construction of flavocytochrome <Emphasis Type="Italic">b</Emphasis><Subscript>2</Subscript>-overproducing strains of the thermotolerant methylotrophic yeast <Emphasis Type="Italic">Hansenula polymorpha</Emphasis> (<Emphasis Type="Italic">Pichia angusta</Emphasis>)

L-Lactate cytochrome c oxidoreductase (flavocytochrome b ₂, FC b ₂) from the thermotolerant methylotrophic yeast Hansenula polymorpha (Pichia angusta) is, unlike the enzyme form baker’s yeast, a thermostable enzyme potentially important for bioanalytical technologies for highly selective assays of L-lactate in biological fluids and foods. This paper describes the construction of flavocytochrome b ₂ producers with over-expression of the H. polymorpha CYB2 gene, encoding FC b ₂. The HpCYB2 gene under the control of the strong H. polymorpha alcohol oxidase promoter in a plasmid for multicopy integration was transformed into the recipient strain H. polymorpha C-105 (grc1 catX), impaired in glucose repression and devoid of catalase activity. A method was developed for preliminary screening of the transformants with increased FC b ₂ activity in permeabilized yeast cells. The optimal cultivation conditions providing for the maximal yield of the target enzyme were found. The constructed strain is a promising FC b ₂ producer characterized by a sixfold increased (to 3 μmol min^?1 mg^?1 protein in cell-free extract) activity of the enzyme.     

Biological activity of new flavonoid from <Emphasis Type="Italic">Hieracium pilosella</Emphasis> L.

Hieracium pilosella L. (Asteraceae) is a well-known plant used in ethno-medicine as its inflorescences are particularly rich in beneficial polyphenolics. This research aimed to elucidate the structure of a new flavone glycoside isolated from the inflorescences of Hieracium pilosella and evaluate its antioxidant, antimicrobial and antiproliferative activities. The chromatographic methods were successfully applied to isolate the new flavonoid. Its structure was determined by subsequent UV, NMR and MS experiments and identified as isoetin 4′-O-β-D-glucopyranoside. Free radical scavenging capacity was examined by measuring the scavenging activity of the new isoetin derivative on 2,2-diphenyl-1-picrylhydrazyl (DPPH). The compound was also screened for spectrum of antimicrobial activity using the agar well diffusion method. Minimum inhibitory concentration (MIC) for Pseudomonas aeruginosa ATCC 9027 was performed by the micro-dilution broth method. The antiproliferative effect of tested glycoside was assessed in two human tumor cell lines derived from lung (A549) and colon (HT-29) carcinoma and cell proliferation was determined by means of MTT method. The tested compound showed high antiradical activity, reducing the DPPH? with EC₅₀ 7.9 μM (3.7 µg/ml) and exhibited narrow antimicrobial spectrum among tested microorganisms. The compound was active against Pseudomonas aeruginosa ATCC 9027 (MIC 125 μg/ml) which is prone to causing infections that are difficult to treat due to it developing extremely rapid antibiotic resistance. In the antiproliferative studies, cell proliferation of the colon (HT-29) carcinoma cell line was significantly decreased after exposure to the compound. The results indicate that isoetin 4′-O-β-D-glucopyranoside possesses antioxidant capacity and very promising antibacterial activity and could have uses as an effective antipseudomonal agent as well a antiproliferative agent.     

Genetics and fine mapping of a yellow-green leaf gene (<Emphasis Type="Italic">ygl</Emphasis>-<Emphasis Type="Italic">1</Emphasis>) in cabbage (<Emphasis Type="Italic">Brassica oleracea</Emphasis> var. <Emphasis Type="Italic">capitata</Emphasis> L.)

Cabbage (Brassica oleracea var. capitata L.) is one of the most popular cultivated vegetables worldwide. Cabbage has rich phenotypic diversity, including plant height, head shape, head color, leaf shape and leaf color. Leaf color plays an important role in cabbage growth and development. At present, there are few reports on fine mapping of leaf color mutants in B. oleracea. In this study, a naturally occurring yellow-green leaf cabbage mutant (YL-1), derived from the self-pollinated progenies of the hybrid ‘Hosom’, was used for inheritance analysis and gene mapping. Segregation populations including F₂ and BC₁ were generated from the cross of two inbred lines, YL-1 and 01–20. Genetic analysis with the F₂ and BC₁ populations demonstrated that the yellow-green leaf color was controlled by a single recessive nuclear gene, ygl-1. Insertion–deletion (InDel) markers, designed based on the parental re-sequencing data, were used for the preliminary mapping with BSA (bulked segregant analysis) method. A genetic map constructed with 15 InDels indicated that ygl-1 was located on chromosome C01. The ygl-1 gene is flanked by InDel markers ID2 and M8, with genetic distances of 0.4 cM and 0.35 cM, respectively. The interval distance between two markers is 167 kb. Thus, it enables us to locate the ygl-1 gene for the first time in B. oleracea. This study lays the foundation for candidate gene prediction and ygl-1gene cloning.     

<Emphasis Type="Italic">In vitro</Emphasis> cytotoxicity of an endophytic fungus isolated from<Emphasis Type="Italic">Nothapodytes foetida</Emphasis>

An attempt has been made to assessin vitro cytotoxicity of an endophytic fungus fromNothapodytes foetida. Various human cancer cell lines (liver HEP-2, lung A-549, ovary OVAR-5, prostate PC-3, cervix Hela, colon HCT-15, oral cell line KB, CNS SNB-78, were used.In vitro cytotoxicity of camptothecin (CPT) isolated from the fungus was done where OVAR-5 cell line showed maximum inhibition and HEP-2 cell line was least sensitive with this compound.In vitro cytotoxicity of fractions/extracts from endophyte was carried out where ethyl acetate fraction showed sufficient growth inhibition against all the cell lines.     

Bacterial metabolites from intra- and inter-species influencing thermotolerance: the case of <Emphasis Type="Italic">Bacillus cereus</Emphasis> and <Emphasis Type="Italic">Geobacillus stearothermophilus</Emphasis>

Bacterial metabolites with communicative functions could provide protection against stress conditions to members of the same species. Yet, information remains limited about protection provided by metabolites in Bacillus cereus and inter-species. This study investigated the effect of extracellular compounds derived from heat shocked (HS) and non-HS cultures of B. cereus and Geobacillus stearothermophilus on the thermotolerance of non-HS vegetative and sporulating B. cereus. Cultures of B. cereus and G. stearothermophilus were subjected to HS (42 or 65 °C respectively for 30 min) or non-HS treatments. Cells and supernatants were separated, mixed in a combined array, and then exposed to 50 °C for 60 min and viable cells determined. For spores, D values (85 and 95 °C) were evaluated after 120 h. In most cases, supernatants from HS B. cereus cultures added to non-HS B. cereus cells caused their thermotolerance to increase (D ₅₀ 12.2–51.9) when compared to supernatants from non-HS cultures (D ₅₀ 7.4–21.7). While the addition of supernatants from HS and non-HS G. stearothermophilus cultures caused the thermotolerance of non-HS cells from B. cereus to decrease initially (D ₅₀ 3.7–7.1), a subsequent increase was detected in most cases (D ₅₀ 18–97.7). In most cases, supernatants from sporulating G. stearothermophilus added to sporulating cells of B. cereus caused the thermotolerance of B. cereus 4810 spores to decline, whereas that of B. cereus 14579 increased. This study clearly shows that metabolites in supernatants from either the same or different species (such as G. stearothermophilus) influence the thermotolerance of B. cereus.