诱导茄链格孢菌分生孢子形成的新技术

本文报道诱导茄链格孢菌(Alternaria solani)分生孢子形成的一种新技术。生长在马铃薯-葡萄糖-琼脂(PDA)上两天的茄链格孢菌琼脂块移接到玉米培养基上,置于日光灯下照射,诱发分生孢子梗生长。然后,再放在18℃下黑暗培养。12小时后,在菌丝块表面有大量的分生孢子形成。成熟的茄链格孢菌分生孢子用蒸馏水洗脱。     

Improving the robustness of a low-cost insect cell medium for baculovirus biopesticides production, via hydrolysate streamlining using a tube bioreactor-based statistical optimization routine

A critical component of an in vitro production process for baculovirus biopesticides is a growth medium that is efficacious, robust, and inexpensive. An in‐house low‐cost serum‐free medium, VPM3, has been shown to be very promising in supporting Helicoverpa armigera nucleopolyhedrovirus (HaSNPV) production in H. zea insect cell suspension cultures, for use as a biopesticide against the Heliothine pest complex. However, VPM3 is composed of a significant number of undefined components, including five different protein hydrolysates, which introduce a challenging lot‐to‐lot variability to the production process. In this study, an intensive statistical optimization routine was employed to reduce the number of protein hydrolysates in VPM3 medium. Nearly 300 runs (including replicates) were conducted with great efficiency by using 50 mL TubeSpin® bioreactors to propagate insect cell suspension cultures. Fractional factorial experiments were first used to determine the most important of the five default protein hydrolysates, and to screen for seven potential substitutes for the default meat peptone, Primatone RL. Validation studies informed by the screening tests showed that promising alternative media could be formulated based on just two protein hydrolysates, in particular the YST‐AMP (Yeast Extract and Amyl Meat Peptone) and YST‐POT (Yeast Extract and Lucratone Potato Peptone) combinations. The YST‐AMP (meat‐based) and YST‐POT (meat‐free) variants of VPM3 were optimized using response surface methodology, and were shown to be just as good as the default VPM3 and the commercial Sf‐900 II media in supporting baculovirus yields, hence providing a means toward a more reproducible and scalable production process for HaSNPV biopesticides. © 2012 American Institute of Chemical Engineers Biotechnol. Prog.,, 2012     

Similarities and differences between the responses induced in human phagocytes through activation of the medium chain fatty acid receptor GPR84 and the short chain fatty acid receptor FFA2R

GPR84 is a recently de-orphanized member of the G-protein coupled receptor (GPCR) family recognizing medium chain fatty acids, and has been suggested to play important roles in inflammation. Due to the lack of potent and selective GPR84 ligands, the basic knowledge related to GPR84 functions is very limited. In this study, we have characterized the GPR84 activation profile and regulation mechanism in human phagocytes, using two recently developed small molecules that specifically target GPR84 agonistically (ZQ16) and antagonistically (GLPG1205), respectively. Compared to our earlier characterization of the short chain fatty acid receptor FFA2R which is functionally expressed in neutrophils but not in monocytes, GPR84 is expressed in both cell types and in monocyte-derived macrophages. In neutrophils, the GPR84 agonist had an activation profile very similar to that of FFA2R. The GPR84-mediated superoxide release was low in naïve cells, but the response could be significantly primed by TNFα and by the actin cytoskeleton disrupting agent Latrunculin A. Similar to that of FFA2R, a desensitization mechanism bypassing the actin cytoskeleton was utilized by GPR84. All ZQ16-mediated cellular responses were sensitive to GLPG1205, confirming the GPR84-dependency. Finally, our data of in vivo transmigrated tissue neutrophils indicate that both GPR84 and FFA2R are involved in neutrophil recruitment processes in vivo.In summary, we show functional similarities but also some important differences between GPR84 and FFA2R in human phagocytes, thus providing some mechanistic insights into GPR84 regulation in blood neutrophils and cells recruited to an aseptic inflammatory site in vivo.     

Effect of cyanobacterial extracellular products on high-frequency in vitro induction and elongation of Gossypium hirsutum L organs through shoot apex explants

The challenging task of bringing high efficiency transformed plants attracts lot of attention in recent times. In search for this, there have been many attempts made using, different techniques like tissue culture and plant breeding methods. Here we report a suitable alternative facile route, where cyanobacterial extracellular products are utilized as growth regulators and its performance validated on Gossypium hirsutum L. MS medium is tested with cyanobacterial extracellular products of Nostoc ellipsosporum, Dolichospermum flos-aquae and Oscillatoria acuminata .Our best results show that the addition of O. acuminata extracellular product with plant growth hormones gives the excellent induction and elongation in cotton. In addition to this, the multiple shoot was obtained on MS medium fortified with 1.0 mg L^?1 BA with 8% O. acuminata and 1.5 mg L^?1 TDZ with 12% O. acuminata. High frequency of shoot elongation supplemented with MS medium, iP 2.5 mg L^?1 and 16% O. acuminata and root production MS medium fortified with 12% O. acuminata best responsible for regeneration in cotton plants. The rooted plants were hardened and transferred to soil with 90% survival rate.     

Importance of some factors on the dimorphism of Candida albicans

The passage between the yeast and mycelial forms of Candida albicans B 311-10 was studied by using the minimal syntehtic medium of Shepherd et al. [19] modified without biotin and with low glucose concentrations. It was observed that biotin, aminoacids and particularly pH are not important factors in the dimorphism of C. albicans. The only factor of notable importance in the passage of yeast form to mycelial form in C. albicans was glucose concentration.     

Phospholipase D1 downregulation by α-synuclein: Implications for neurodegeneration in Parkinson's disease

We have previously shown that phospholipase D (PLD) pathways have a role in neuronal degeneration; in particular, we found that PLD activation is associated with synaptic injury induced by oxidative stress. In the present study, we investigated the effect of α-synuclein (α-syn) overexpression on PLD signaling. Wild Type (WT) α-syn was found to trigger the inhibition of PLD1 expression as well as a decrease in ERK1/2 phosphorylation and expression levels. Moreover, ERK1/2 subcellular localization was shown to be modulated by WT α-syn in a PLD1-dependent manner. Indeed, PLD1 inhibition was found to alter the neurofilament network and F-actin distribution regardless of the presence of WT α-syn. In line with this, neuroblastoma cells expressing WT α-syn exhibited a degenerative-like phenotype characterized by a marked reduction in neurofilament light subunit (NFL) expression and the rearrangement of the F-actin organization, compared with either the untransfected or the empty vector-transfected cells. The gain of function of PLD1 through the overexpression of its active form had the effect of restoring NFL expression in WT α-syn neurons. Taken together, our findings reveal an unforeseen role for α-syn in PLD regulation: PLD1 downregulation may constitute an early mechanism in the initial stages of WT α-syn-triggered neurodegeneration.     

Medium components and culture conditions affect the thermotolerance of aerial conidia of fungal biocontrol agent Beauveria bassiana

AIMS: To produce more thermotolerable conidia of Beauveria bassiana, a well-known fungal biocontrol agent, by optimizing the medium components and culture conditions. METHODS AND RESULTS: The conidia produced on media including 0.5-6% glucose, sucrose or starch as carbon source and 50-300-microg ml(-1) Cu2+, Zn2+, Mn2+ or Fe3+ as additive to Sabouraud dextrose medium at 15-30 degrees C, pH 4-8 or KCl-adjusted water availabilities were exposed to 30-min wet heat stress at 48 degrees C. The medium components for conidial production with greatly enhanced thermotolerance included 4% glucose as optimum or 1% starch as alternative for the carbon source and < or =50-microg ml(-1) Mn2+ for the metal additive. The culture conditions were optimized as 25 degrees C and pH 5-6. Conidial thermotolerance decreased remarkably when sucrose and Fe3+ or Cu2+ were used in the cultures, but altered slightly when 50-200-microg ml(-1) Zn2+ were included. CONCLUSIONS: The tolerance of B. bassiana conidia to the thermal stress was significantly affected by the medium composition and culture conditions under which the conidia were produced. SIGNIFICANCE AND IMPACT OF THE STUDY: Proper treatment of small grains as mass production substrates for more glucose release and supplement of glucose or 50-microg ml(-1) Mn2+ are possible means to enhancing conidial thermotolerance and field persistence for improved insect control.     

Stochastic exposure to sub-lethal high temperature enhances exopolysaccharides (EPS) excretion and improves Bifidobacterium bifidum cell survival to freeze–drying

Exposure of microbial cells to sub-lethal stresses is known to increase cell robustness. In this work, a two-compartment bioreactor in which microbial cells are stochastically exposed to sub-lethal temperature stresses has been used in order to investigate the response of the stress sensitive Bifidobacterium bifidum THT 0101 to downstream processing operations. A stochastic model validated by residence time distribution experiments has shown that in the heat-shock configuration, a two-compartment bioreactor (TCB) allows the exposure of microbial cells to sub-lethal temperature of 42 °C for a duration comprised between 100 and 300 s. This exposure resulted in a significant increase of cell resistance to freeze–drying by comparison with cells cultivated in conventional bioreactors or in the TCB in the cold shock mode (CS-TCB). The mechanism behind this robustness seems to be related with the coating of microbial cells with exopolysaccharide (EPS), as assessed by the change of the zeta potential and the presence of higher EPS concentration after heat shock. Conditioning of Bifidobacteria on the basis of the heat shock technique is interesting from the practical and economical point of view since this strategy can be directly implemented in the bioreactor during stationary phase preceding cell recovery and freeze–drying.     

Microresonator mass sensors for detection of Bacillus anthracis Sterne spores in air and water

Towards the goal of developing a real-time monitoring device for microorganisms, we demonstrate the use of microcantilevers as resonant mass sensors for detection of Bacillus anthracis Sterne spores in air and liquid. The detection scheme was based on measuring resonant frequency decrease driven by thermally induced oscillations, as a result of the added mass of the spores with the use of a laser Doppler vibrometer (LDV). Viscous effects were investigated by comparing measurements in air and deionized (DI) water along with theoretical values. Moreover, biological experiments were performed which involved suspending spores onto the cantilevers and performing mass detection in air and water. For detection of spores in water, the cantilevers were functionalized with antibodies in order to fix the spores onto the surface. We demonstrate that as few as 50 spores on the cantilever can be detected in water using the thermal noise as excitation source. Measurement sensitivity of 9.23 Hz/fg for air and 0.1 Hz/fg for water were obtained. These measurements were compared with theoretical values and sources of improvement in cantilever sensitivity in a viscous medium were also discussed. It is expected that by driving the cantilevers and using higher order modes, detection of a single spore in liquids should be achievable.