Breeding of mycoparasitic Trichoderma strains for heavy metal resistance

AIMS: This study was designed to investigate the effects of 10 heavy metals on the in vitro activities of beta-glucosidase, cellobiohydrolase, beta-xylosidase and endoxylanase enzymes for six strains of Trichoderma, and to isolate and characterize heavy metal-resistant mutants. METHODS AND RESULTS: At a concentration of 1 mmol, only mercury showed significant inhibitory effects on the in vitro enzyme activities; in all other cases, the enzymes remained active. A total of 177 heavy metal-resistant mutants were isolated and tested for cross-resistance to other heavy metals. Some mutants were effective antagonists of Fusarium, Pythium and Rhizoctonia strains, even on media containing the respective heavy metals. CONCLUSION: Trichoderma strains could be developed as biocontrol agents that are effective against plant pathogenic fungi, even under heavy metal stress. SIGNIFICANCE AND IMPACT OF THE STUDY: Trichoderma mutants resistant to heavy metals might be of value for use with heavy metal-containing pesticides, as part of an integrated plant protection system.     

Complex homogeneous and heterogeneous fluorescence anisotropy decays: enhancing analysis accuracy

In biological macromolecules, fluorophores often exhibit multiple depolarizing motions that require multiple lifetimes and rotational relaxation times to define fluorescence intensity and anisotropy decays. The related analysis of time-correlated single-photon counting data becomes uncertain due to the multitude of decay parameters and numerical sensitivity to deconvolution of the instrument response function (IRF) via discretization of integrals. By using simulations we show that improved discretizations based on quadratic and cubic local approximations of the IRF yield more accurate estimation of short rotational relaxation times and lifetimes than the commonly used Grinvald-Steinberg discretization, which in turn appears more reliable than two discretizations based on linear local approximations of the IRF. In addition, our simulation suggests that cubic approximation is the most advantageous in discriminating complex heterogeneous and homogeneous anisotropy decay. We show that among three different information criteria, the Akaike information criterion is best suited for detection of heterogeneity in rotational relaxation times. It is capable of detecting heterogeneity even when anisotropy decay appears homogeneous within statistical errors of estimation.     

Genistein对大鼠垂体前叶细胞增殖的抑制作用

应用细胞培养、^3H-TdR掺入、流式细胞和电镜技术,观察酪氨酸蛋白激酶（PTK）抑制剂genistein对正常大鼠垂体前叶细胞和垂体瘤细胞株AtT-20增殖的影响,并探讨其可能的机制。结果显示：genistein作用48h后可明显抑制正常大鼠垂体前叶细胞和垂体瘤细胞株AtT-20增殖。流式细胞仪检测发现,50和100μmol/L genistein可将AtT-20细胞阻断于G0／G1期及G2／M期,并出现凋亡峰,凋亡率分别灰19.9％和36.4％。电镜照片显示有凋亡细胞。结果表明,PTK抑制剂可以明显抑制正常大鼠垂体前叶细胞和垂体瘤细胞株AtT-20的殖,并诱导细胞凋亡,说明PTK活性对细胞增殖和分化有重要作用。     

非NMDA受体参与双相呼气和吸气神经元电活动的调节

在新生大鼠延髓脑片上同步记录舌下神经根和双相呼气神经元／吸气神经元单位的放电活动,并在灌流的改良Kredbs液中先后加以非NMDA受体的激动剂KA和拮抗剂DNQX,观察对神经元单位放电的影响,以进一步探讨非NMDA受体在对双相呼气神经元之间交互兴奋和吸气神经元兴奋性突触输入中的作用,结果表明,使用非NMDA受体激动剂KA以后,双相呼气神经元的放电频率和蜂频率都明显增大,吸气神经元中期放电的频率和非NMDA受体激动剂KA以后,双相呼气神经元的放电频率和峰频率都明显增大,吸气神经元中期放电的频率和峰频率也显著增大,而早期和晚期放电的频率无明显改变,用相应拮抗剂以后,上述效应明显被抑制,结果提示,非NMDA受体参与了双相呼气神经元之间的交互兴奋作用,并且也介导了吸气神经元的兴奋性突触输入／     

Platelet-derived growth factor C (PDGF-C), a novel growth factor that binds to PDGF alpha and beta receptor

We have characterized platelet-derived growth factor (PDGF) C, a novel growth factor belonging to the PDGF family. PDGF-C is a multidomain protein with the N-terminal region homologous to the extracellular CUB domain of neuropilin-1, and the C-terminal region consists of a growth factor domain (GFD) with homology to vascular endothelial growth factor (25%) and PDGF A-chain (23%). A serum-sensitive cleavage site between the two domains allows release of the GFD from the CUB domain. Competition binding and immunoprecipitation studies on cells bearing both PDGF alpha and beta receptors reveal a high affinity binding of recombinant GFD (PDGF-CC) to PDGF receptor-alpha homodimers and PDGF receptor-alpha/beta heterodimers. PDGF-CC exhibits greater mitogenic potency than PDGF-AA and comparable or greater mitogenic activity than PDGF-AB and PDGF-BB on several mesenchymal cell types. Analysis of PDGF-CC in vivo in a diabetic mouse model of delayed wound healing showed that PDGF-CC significantly enhanced repair of a full-thickness skin excision. Together, these studies describe a third member of the PDGF family (PDGF-C) as a potent mitogen for cells of mesenchymal origin in in vitro and in vivo systems with a binding pattern similar to PDGF-AB.     

Agrobacterium rhizogenes-mediated transformation and the regeneration of transformants in Alhagi pseudalhagi]

The regenerated shoot segments of Alhagi pseudalhagi were sliced and infected with Agrobacterium rhizogenes strain A4. The hairy roots and transformed calli were obtained through selection on hormone free MS medium. The transformants were cultured on MS medium with 2 mg/L 2,4-dichlorophenoxy acetic acid (2,4-D) and 0.5-1 mg/L 6-benzylaminopurine (6-BA) to induce calli. 3 mg/L 6-BA and 0.5 mg/L naphthalene acetic acid (NAA) were applied for shoot differentiation. Shoots were planted on MS medium with 2 mg/L indole-3-butyric acid (IBA) and produced roots. Opine analysis proved the integration and expression of T-DNA in over 95% hairy roots, 75% transformed calli and transformed plantlets respectively. The 81% hairy root cells had normal chromosome numbers (2n = 18). The alterations of chromosome number were observed. After one year of subculturing, the regeneration ability of transformants was maintained.     

Early Archean planktonic mode of life: Implications from fluid dynamics of lenticular microfossils

Lenticular, and commonly flanged, microfossils in 3.0–3.4 Ga sedimentary deposits in Western Australia and South Africa are unusually large (20–80 μm across), robust, and widespread in space and time. To gain insight into the ecology of these organisms, we performed simulations of fluid dynamics of virtual cells mimicking lenticular forms of variable sizes, oblateness, flange presence, and flange thickness. Results demonstrate that (a) the flange reduces sedimentation velocity, (b) this flange function works more effectively in larger cells, and (c) modest oblateness lowers sedimentation rate. These observations support interpretations that the lenticular microbes were planktonic—a lifestyle that could have been advantageous in an early Earth harsh environment including violent volcanic activities, repeated asteroid impacts, and relatively high UV‐radiation. Although the robustness of these organisms could have provided additional protection on the early Earth, this architecture may have impeded a planktonic lifestyle by increasing cell density. However, our data suggest that this disadvantage could have been compensated by enlargement of cell volume, which could have enhanced the ability of the flange to slow sedimentation rate, especially if coupled with vacuolation. The results of this simulation study may help to explain the unique morphology and unusually large size of these Archean microfossils.     

Development of a loop‐mediated isothermal amplification assay for rapid and sensitive detection of Sporisorium scitamineum in sugarcane

Smut disease caused by Sporisorium scitamineum is one of the most destructive sugarcane diseases worldwide. The pathogen spreads primarily through infected sugarcane setts, and hence, the use of disease‐free planting materials is essential for preventing disease development in the field. In this study, a species‐specific loop‐mediated isothermal amplification (LAMP) assay was developed for rapid and accurate detection of S. scitamineum. Based on the differences in internal transcribed spacer (ITS) sequences of S. scitamineum, a set of four species‐specific primers, F3, B3, FIP and BIP, were designed by using a panel of fungal and bacterial species as controls. After optimization of the reaction conditions, the detection limit of LAMP assay was about 2 fg of the S. scitamineum genomic DNA in 25 µL reaction solution, 100‐fold lower than that of conventional polymerase chain reaction. The assay showed high specificity to discriminate all S. scitamineum isolates from nine other fungal and bacterial pathogens. The LAMP assay also detected smut infection from young sugarcane leaves with no visible smut‐disease symptoms. The findings from this study provide a simple, highly sensitive, rapid and reliable technique for early detection of S. scitamineum, which may be useful for sugarcane quarantine and production of smut‐free seedcanes. This is the first report of LAMP‐based assay for the detection of S. scitamineum in sugarcane.