The first study on the effects of microcystin-RR on gene expression profiles of antioxidant enzymes and heat shock protein-70 in Synechocystis sp PCC6803

Microcystins are heptapeptide toxins produced by cyanobacteria. Microcystin-RR(MC-RR) is a common variant among the 80 variants identified so far. There have been many investigations documenting the toxic effects of microcystins on animals and higher plants, but little is known on the toxic effects of microcystins on algae, especially at molecular level. We studied the effects of MC-RR on gene expression profile of a few antioxidant enzymes and heat shock protein-70 (Hsp70) in Synechocystis sp. PCC6803. After two days post-exposure, a high dose toxin (5 mg/l, about 4.8 x 10(-3) mM) significantly increased expression levels of the genes gpx1, sodB, katG, acnB, gamma-TMTand dnaK2, while a relatively low dose toxin (1 mg/l, about 9.63 x 10(-4) mM) induced a moderate and slow increase of gene expression. Our results indicate that MC-RR could induce the oxidative stress in Synechocystis sp. PCC6803 and the increase in gene expression of antioxidant enzymes and Hsp70 might protect the organism from the oxidative damage. in addition, cell aggregation was observed during the early period of exposure, which might be a specific oxidative stress reaction to MC-RR. (C) 2008 Elsevier Ltd. All rights reserved.

Kinetic analysis of interaction between lipopolysaccharide and biomolecules

Lipopolysaccharide ( LPS) is a major component of the outer membrane of all gram-negative bacteria. It is a heat-resistant toxin which can cause toxic shock in animals. LPS interacts with some biomolecules and triggers its toxic reaction. In this study, the interaction between LPS from Salmonella Minnesota and some biomolecules using syrface okasnib resibabce ( SPR) biosensor. biomolecules were imobilized on CM5 sensor-chip suing amion coupling method and LPS was injected over the immobilized surfaces.

Toxin-screening and identification of bacteria isolated from highly toxic marine gastropod Nassarius semiplicatus

Bacteria isolated from a highly toxic sample of gastropod Nassarius semiplicatus in Lianyungang, Jiangsu Province in July 2007, were studied to probe into the relationship between bacteria and toxicity of nassariid gastropod. The toxicity of the gastropod sample was 2 x 10(2) mouse unit (MU) Per gram Of tissue (wet weight). High concentration of tetrodotoxin (TTX) and its analogues (TTXs) were found in the digestive gland and muscle of the gastropod, using high performance liquid chromatography coupled with mass chromatography (LC-MS). Bacterial strains isolated from the digestive gland were cultured and screened for TTX with a competitive ELISA method. Tetrodotoxin was detected in a proportion of bacterial strains, but the toxin content was low. Partial 16S ribosomal DNA (rDNA) of the TTX-producing strains was then sequenced and compared with those published in the GenBank to tentatively identify the toxic strains. It was found that most of the toxic strains were closely affiliated with genus Vibrio, and the others were related to genus Shewanella, Marinomonas, Tenacibaculum and Aeromonas. These findings suggest that tetrodotoxin-producing bacteria might play an important role in tetrodotoxin accumulation/production in N. semiplicatus. (C) 2008 Elsevier Ltd. All rights reserved.

Thermal Decomposition of 1,2-Dichloroethane in a Single-Pulse Shock Tube

在单脉冲激波管上,研究了1,2-二氯乙烷的热裂解.实验的激波条件为:温度区间1020 K＜T＜1190 K, 压力: P=0.12 MPa,实验时间τ=0.5 ms;实验气体为1,2-二氯乙烷稀释于Ar气中(3.95 mmol/L).以4-甲基-1-环己烯作为对比速率法实验的内标物,用4-甲基-1-环己烯开环反应的速率常数k=1015.3exp(-33400/T) s-1,以及从其产物的浓度推定出实验温度.经激波加热后的实验气体的终产物用气相色谱分析出主要成分为C2H3Cl,指示出主要反应通道为β消去反应.如把所有产物C2H3Cl都归于β消去反应,则可推定出表观之反应速率常数k1a=5.0×1013exp(-30000/T) s-1.对于由C-Cl键断键反应引发的链反应的可能影响做了分析研究.用了一种简便分析可推知在实验的温度范围内的低端(1020 K)链反应的影响可以忽略,而在其高端(1190 K)链反应将给出10%的终产物C2H3Cl的附加浓度,获得真实的β消去反应速率常数则必须把这部分予以扣除.经过这样的校正之后,最后得到CH2ClCH2Clβ消去反应速率常数为k1c=2.3×1013exp(-29200/T) s-1.

Effects of pure microcystin-LR on the transcription of immune related genes and heat shock proteins in larval stage of zebrafish (Danio rerio)

Microcystins (MCs) are cyanobacterial toxins in water blooms that have received increasing attention as a public biohazard for human and animal health. Previous studies were mainly focused on the toxic effects on adult fish, rather than juvenile or larvae, and the response of fish immune system were usually neglected. This paper presents the first data of the effects of microcystin-LR (MC-LR) on transcription of several genes essential for early lymphoid development (Rag1, Rag2, Ikaros, GATA1, Lck and TCR alpha) and heat shock proteins (HSP90, HSP70, HSP60, HSP27) in zebrafish larvae. Relative changes of mRNA transcription were analyzed by real time PCR. The transcription of Rag1, Rag2, Ikaros, GATA1, Lck and TCR alpha were up-regulated when following exposure to 800 mu g/L MC-LR, which may indicate that specific lymphocytes differentiation and TCR/lg arrangement are induced to counteract the toxic effects of MC-LR. It was also interesting to note the dramatically increased transcription of HSP90. HSP70, HSP60 and HSP27, which may indicate their important roles as molecular chaperones under oxidative stress. (C) 2009 Elsevier B.V. All rights reserved.

Bioaccumulation of the hepatotoxic microcystins in various organs of a freshwater snail from a subtropical Chinese lake, Taihu lake, with dense toxic microcystis blooms

In this paper, we describe the seasonal dynamics of three common microcystins (MCs MC-RR, MC-YR, and MC-LR) in the whole body, hepatopancreas, intestine, gonad, foot, remaining tissue, and offspring of a freshwater snail, Bellamya aeruginosa, from Gonghu Bay of Lake Taihu, China, where dense toxic Microcystis blooms occur in the warm seasons. Microcystins were determined by liquid chromatography electrospray ionization mass spectrum. Microcystin (MC-RR + MC-YR + MC-LR) content of the offspring and gonad showed high positive correlation, indicating that microcystins could transfer from adult females to their young with physiological connection. This study is the first to report the presence of microcystins in the offspring of the adult snail. The majority of the toxins were present in the intestine (53.6%) and hepatopancreas (29.9%), whereas other tissues contained only 16.5%. If intestines are excluded, up to 64.3% of the toxin burden was allocated in the hepatopancreas. The microcystin content in the intestine, hepatopancreas, and gonad were correlated with the biomass of Microcystis and intracellular and extracellular toxins. Of the analyzed foot samples, 18.2% were above the tolerable daily microcystin intake recommended by the World Health Organization (WHO) for human consumption. This result indicates that public health warnings regarding human ingestion of snails from Taihu Lake are warranted. In addition, further studies are needed to evaluate the occurrence by Microcystis in relation to spatial and temporal changes in water quality.

In situ studies on the bioaccumulation of microcystins in the phytoplanktivorous silver carp (Hypophthalmichthys molitrix) stocked in Lake Taihu with dense toxic Microcystis blooms

The phytoplanktivorous silver carp is an important biomanipulation fish to control cyanobacterial blooms and is also a food fish with the greatest production in China. The accumulation of the hepatotoxic microcystins (MCs) determined by LC-MS in various organs of silver carp was studied monthly in Lake Taihu dominated by toxic Microcystis aeruginosa. Average recoveries of spiked fish samples were 78% for MC-RR and 81% for MC-LR. The highest content of MCs was found in the intestine (97.48 mu g g(-1) DW), followed by liver (6.84 mu g g(-1) DW), kidney (4.8 8 mu g g(-1) DW) and blood (1.54 mu g g(-1) DW), and the annual mean MC content was in the order of intestine > liver > kidney > blood > muscle > spleen > gallbladder > gill. Silver carp could effectively ingest toxic Microcystis cells (up to 84.4% of total phytoplankton in gut contents), but showed fast growth (from 141 g to 1759 g in I year in mean weight). Silver carp accumulated less microcystins in liver than other animals in the same site or other fish from different water bodies at similar level of toxin ingestion. There was possible inhibition of the transportation of the most toxic MC-LR across the gutwall. Muscle of silver carp in Lake Taihu should not be consumed during period of dense Microcystis blooms while viscera were risky for consumption in more months. (c) 2006 Elsevier B.V. All rights reserved.

Analysis of paralytic shellfish toxins in Aphanizomenon DC-1 from Lake Dianchi, China

Lake Dianchi is in Yunnan Province in southwestern China. In recent years, significant cyanobacterial blooms have occurred in this lake nearly every year because of eutrophication. Monitoring data for the past 5 years acquired by our research group showed that phytoplankton composition alternated between species of Microcystis sp. during warm seasons and those of Aphanizomenon sp. during cool seasons. In March 2003, when phytoplankton composition was highly dominated by Aphanizomenon sp., samples were taken from the lake for toxin detection and immediate strain isolation. A mouse bioassay with extracts from the lyophilized field material showed obvious intoxication from paralytic shellfish poisons (PSPs), and all mice died within 30 min. Further analysis of both field and isolated algal strain Aphanizomenon DC-1 by the postcolumn HPLC-FLD method confirmed its PSP-producing ability The analogues found in the extracts from the field material were neoSTX, dcSTX, and dcGTX3, with contents of 2.279, 1.135, and 0.547 ng/mg DW, respectively. Under laboratory culture condition, toxin content in the Aphanizomenon strain DC-1 varied greatly during different growth phases, with two peaks: in the early-exponential and late-stationary growth phases. When the culture grew at a relatively high rate during the mid- to late-exponential growth phase, toxin content declined gradually. Moreover, the types of toxin in the DC-1 strain varied greatly during a single culture cycle. The HPLC results showed that dcSTX was the only toxin isomer detected throughout the culture period, and its level remained stable. On the other hand, dcGTX2 and GTX4 were the major toxins during the early-exponential and stationary phases, respectively. This article presents the first data on the identification and detection of paralytic shellfish toxins from cyanobacteria in Lake Dianchi. As far as we know, this is also the first report of this type of toxin in inland water bodies in China. Our study indicates the threat associated with PSP toxins in Lake Dianchi and suggests that necessary measures and programs for control are urgently needed to prevent the spread of toxic cyanobacterial blooms. (c) 2006 Wiley Periodicals, Inc.

Effects of nitrogen forms on the production of cyanobacterial toxin microcystin-LR by an isolated Microcystis aeruginosa

A cyanobacterial strain, which produced high content of microcystin-LR (MC-LR) but no rnicrocystin-RR (MC-RR), was isolated from the hypertrophic Dianchi Lake in China and identified as Microcystis aeruginosa DC-1. Effects of nitrogen containing chemicals and trace elements on the growth and the production of MC-LR by this strain were Studied. In the presence of bicine, compared with urea and ammonium, nitrate greatly promoted the growth and the production of MC-LR. However, leucine and arginine, which were the constitutional components in the molecular structure of MC-LR or RR, inhibited the production of MC-LR. Iron and silicon up to 10mg/L had little effects on the growth of M. aeruginosa DC-1, but the production of MC-LR was apparently enhanced. Under all conditions studied here, only MC-LR but no RR was detected within the cells of M. aeruginosa DC-1. Thus, chemical forms of nitrogen, rather than the usually concerned the total nitrogen, Lind trace elements played important roles in the production of MC toxins during cyanobacterial blooms.

Responses of antioxidant systems in the hepatocytes of common carp (Cyprinus carpio L.) to the toxicity of microcystin-LR

The freshwater, bloom-forming cyanobacterium (blue-green alga) Microcystis aeruginosa produces a peptide hepatotoxin, which causes the damage of animal liver. Recently, toxic Microcystis blooms frequently occur in the eutrophic Dianchi Lake (300 km(2) and located in the South-Westem of China). Microcystin-LR from Microcystis in Dianchi was isolated and purified by high performance liquid chromatography (HPLC) and its toxicity to mouse and fish liver was studied (Li et al., 2001). In this study, six biochemical parameters (reactive oxygen species, glutathione, superoxide dismutase, catalase, glutathione peroxide and glutathione S-transferase) were determined in common carp hepatocytes when the cells were exposed to 10 mug microcystin-LR per litre. The results showed that reactive oxygen species (ROS) contents increased by more than one-time compared with the control after 6 h exposure to the toxin. In contrast, glutathione (GSH) levels in the hepatocytes exposed to microcystin-LR decreased by 47% compared with the control. The activities of superoxide dismutase (SOD), catalase (CAT) and glutathione peroxide (GSH-Px) increased significantly after 6 h exposure to microcystin-LR, but glutathione S-transferase (GST) activity showed no difference from the control. These results suggested that the toxicity of microcystin-LR caused the increase of ROS contents and the depletion of GSH in hepatocytes exposed to the toxin and these changes led to oxidant shock in hepatocytes. Increases of SOD, CAT and GSH-Px activities revealed that these three kinds of antioxidant enzymes might play important roles in eliminating the excessive ROS. This paper also examined the possible toxicity mechanism of microcystin-LR on the fish hepatocytes and the results were similar to those with mouse hepatocytes. (C) 2003 Elsevier Science Ltd. All rights reserved.

Direct toxicity assessment of toxic chemicals with electrochemical method

Electrochemical measurement of respiratory chain activity is a rapid and reliable screening for the toxicity on microorganisms. Here, we investigated in-vitro effects of toxin on Escherichia coli (E. coli) that was taken as a model microorganism incubated with ferricyanide. The current signal of ferrocyanide effectively amplified by ultramicroelectrode array (UMEA), which was proven to be directly related to the toxicity. Accordingly, a direct toxicity assessment (DTA) based on chronoamperometry was proposed to detect the effect of toxic chemicals on microorganisms. The electrochemical responses to 3,5-dichlorophenol (DCP) under the incubation times revealed that the toxicity reached a stable level at 60 min, and its 50% inhibiting concentration (IC50) was estimated to be 8.0 mg L-1. At 60 min incubation, the IC50 values for KCN and As2O3 in water samples were 4.9 mg L-1 and 18.3 mg L-1, respectively. But the heavy metal ions, such as Cu2+ Pb2+ and Ni2+, showed no obvious toxicity on E. coli.

Temperature measurement of reflected shock wave by using chemical indicator

This report describes a new method for measuring the temperature of the gas behind the reflected shock wave in shock tube, corresponding to the reservoir temperature of a shock tunnel, based on the chemical reaction of small amount of CF4 premixed in the test gas. The final product C2F4 is used as the temperature indicator, which is sampled and detected by a gas chromatography in the experiment. The detected concentration of C2F4 is correlated to the temperature of the reflected shock wave with the initial pressure P-1 and test time tau as parameters in the temperature range 3 300 K < T < 5 600 K, pressure range 5 kPa < P1 <12 kPa and tau similar or equal to 0.4 ms.

Micro-shock waves generated inside a fluid jet impinging on plane wall

An Nd:glass laser pulse (18 ns, 1.38 J) is focused in a tiny area of about 100-mum diam under ambient conditions to produce micro-shock waves. The laser is focused above a planar surface with a typical standoff distance of about 4 mm, The laser energy is focused inside a supersonic circular jet of carbon dioxide gas produced by a nozzle with internal diameter of 2.9 mm and external diameter of 8 mm, Nominal value of the Mach number of the jet is around 2 with the corresponding pressure ratio of 7.5 (stagnation pressure/static pressure at the exit of the nozzle), The interaction process of the micro-shock wave generated inside the supersonic jet with the plane wall is investigated using double-pulse holographic interferometry. A strong surface vortex field with subsequent generation of a side jet propagating outward along the plane wail is observed. The interaction of the micro-shock wave with the cellular structure of the supersonic jet does not seem to influence the near surface features of the flowfield. The development of the coherent structures near the nozzle exit due to the upstream propagation of pressure waves seems to be affected by the outward propagating micro-shock wave. Mach reflection is observed when the micro-shock wave interacts with the plane wall at a standoff distance of 4 mm, The Mach stem is slightly deflected, indicating strong boundary-layer and viscous effects near the wall. The interaction process is also simulated numerically using an axisymmetric transient laminar Navier-Stokes solver. Qualitative agreement between experimental and numerical results is good.

Shock Tube Studies on Recombination Kinetics of Sodium Ion with Electron

The ionization kinetics of sodium diluted in argon is studied in a shock tube, in which the test gas mixture is ionized by a reflected shock wave and subsequently quenched by a strong rarefaction wave. A Langmuir electrostatic probe is used to monitor the variation of the ion number density at the reflection shock wave region. The working state of the probe is in the near fi-ee fall region and a correction for reduction of the probe current due to elastic scattering in the probe sheath is introduced. At the temperature range of 800 to 2600 K and in the ambience of argon gas, the three-body recombination rate coefficient of the sodium ion with electron is determined: 3.43 x 10(-14)T(-3.77) cm(6).s(-1).