Biochemical genetic variation among populations of the greenshell mussel, Perna canaliculus, from New Zealand: preliminary findings

Seven polymorphic and four monomorphic allozyme loci were assayed from nine wild populations and one cultured population of the endemic New Zealand greenshell mussel, Perna canaliculus. Genetic variation was examined to determine the extent of intra-population subdivision and inter-population variability. Five of seven polymorphic loci exhibited significant heterozygote deficiencies compared to Hardy-Weinberg equilibrium. Population F-statistics indicated that significant genetic heterogeneity exists among populations, indicating that there is insufficient gene flow between geographically isolated populations to create panmixia. The observed genetic heterogeneity among populations is best explained by an isolation-by-distance model of gene flow, which is modified by localized hydrographic conditions. These preliminary findings are discussed in the context of the one previous survey of population genetic variation in P. canaliculus and how this information relates to the gene flow of the greenshell mussel in New Zealand, which is often mediated by human transport from the main collection site of Kaitaia to aquaculture sites throughout the country.     

Response of brown mussel, Perna indica, to elevated temperatures in relation to power plant biofouling control

1. 1. Results of a study on lethal and sublethal responses of different size groups of the tropical brown mussel, Perna indica, when exposed to different temperatures are presented.

2. 2. Exposure to a temperature of 38°C showed 100% mortality of 9 mm size group mussels in 120 min.

3. 3. Mortality was dependent on age (size) of the mussels, young ones being more susceptible than older ones.

4. 4. All size groups showed a progressive reduction in physiological activities such as filtration rate, foot activity and byssus thread production when temperature was increased from 30°C.

5. 5. This study suggests that heat treatment is an attractive alternative to chlorination for mussel fouling control in tropical power stations.

     

Over-expression of highly conserved mitochondrial 70-kDa heat-shock protein in the sea anemone Anemonia viridis

Organisms react to cellular stress by inducing the synthesis of heat-shock proteins (Hsp). One such protein is the mitochondrial 70-kDa Hsp (mHsp70). The expression of mHsp70 in organisms that undergo stress in their natural habitat is unknown. We used a biochemical approach enabling to identify an mHsp70 from the sea anemone Anemonia viridis, which is abundant in highly fluctuating marine habitats. Antibodies raised specifically against yeast mHsp70 recognized a 70-kDa protein from A. viridis. We found that A. viridis mHsp70 is constitutively expressed at 22–23 °C, but over-expressed upon exposure to heat shock (31 °C) or to temperature fluctuations, suggesting that mHsp70 plays a significant role in adaptation of sessile marine invertebrates to highly fluctuating environmental conditions. Using an affinity column we were able to obtain a partially purified fraction of this protein. Partial amino acid sequences proved that the purified Hsp70 functions in the mitochondria.     

Isoform‐specific responses of metallothioneins in a marine pollution biomonitor,the green‐lipped mussel Perna viridis,towards different stress stimulations

Metallothioneins (MTs) are commonly used as biomarker for metal pollution assessment in marine ecosystems. Using integrated genomic and proteomic analyses, this study characterized two types of MT isoform in the digestive gland of a common biomonitor, the green‐lipped mussel Perna viridis, towards the challenges of a metal (cadmium; Cd) and a non‐metal oxidant (hydrogen peroxide; H₂O₂) respectively. The two isoforms differed in their deduced protein sequences, with 73 amino acids for MT10‐I and 72 for MT10‐II (a novel type), but both consisted of a high percentage (27.4 to 29.2%) of cysteine. Two‐dimensional gel and Western blot showed that the MT proteins were present in multiple isoform spots, and they were further validated to be MT10‐I and MT10‐II using MS analysis coupled with unrestricted modifications searching. Expression of mRNA revealed that MT10‐I responded promptly to Cd but had a lagged induction to H₂O₂ treatments, while MT10‐II was exclusively induced by Cd treatment over the course of exposure. Expression of the MT proteins also showed a delayed response to H₂O₂, compared to Cd treatments. This study uncovered the potential different functional roles of various MTs isoforms in P. viridis and thus advances the resolution of using MTs as biomarkers in future applications.     

Evidence for two Qa-like quinone binding sites in the reaction centre of Rhodopseudomonas viridis

The oxidation-reduction potential of the iron-quinone electron acceptors in the reaction centre of Rhodopseudomonas viridis has been reinvestigated. In chromatophores treated with o-phenanthroline to remove the secondary acceptor Q_b, two steps were observed in the reduction of the primary electron acceptor Q_a with E_m ≈ − 100 and ≈ − 330 mV. In isolated reaction centres only one step was observed in the reduction of Q_a with E ≈ −150 mV. Reconstitution of the reaction centres with additional menaquinone resulted in an increase in the Q_a EPR signal and reconstitution of the low-potential step in the oxidation-reduction titration. Reconstitution with ubiquinone resulted in the recovery of the secondary quinone Q_b. The addition of ubiquinone did not reconstitute the low-potential step of Q_a reduction, or affect the reconstitution of this step by menaquinone. It is concluded that menaquinone can bind to two sites on the reaction centre. Both have properties of the Q_a site but with different pK values. It is unlikely that either is the same as the Q_b site.     

Ecophysiological aspects of cardiac activity in the subtropical mussel Perna viridis (L.) (Bivalvia: Mytilidae)

Cardiac activity in the subtropical mytilid mussel Perna viridis was monitored to assess the influence of various endogenous and exogenous parameters. Temperature had a positive, significant, effect on heart rate; the Q₁₀ ranged from 2.5 to 2.6. Salinity, over a wide range (15-35‰), had no significant influence on heart rate reflecting the species' euryhalinity. Reduced oxygen tensions, either induced by emersion or seawater hypoxia, elicited rapid, significant, bradycardia. Positive inotropisms (indicative of greater heart output) were evident during seawater hypoxia and may represent an energetically advantageous mechanism of reducing heart rate while increasing stroke volume to maintain haemolymph circulation to organs associated with digestion and basal metabolism. Malnourishment for 14 days did not affect heart rate. The heart rate of larger mussels was slower than those of smaller individuals. Gender-based differences in P. viridis' heart rate were not present. No diurnal (24 h) variation in cardiac activity was evident while mussels were exposed to relatively stable, immersed conditions. Heart rate plasticity often reflected the ambient environmental conditions and such a flexible physiological response may partly explain the species' dominance in transitional, polluted harbours.     

假结核耶尔森氏菌中Phd-Doc毒素-抗毒素系统的功能鉴定

【背景】毒素-抗毒素系统在微生物体内广泛存在,在微生物对抗外界不良环境方面发挥重要作用。【目的】以模式细菌假结核耶尔森氏菌(Yersinia pseudotuberculosis,Yptb)为材料,探究其编码的Phd-Doc毒素-抗毒素系统的作用机制和生物学功能。【方法】通过生物信息学方法预测Yptb中编码的Phd-Doc毒素-抗毒素系统,通过毒性分析、基因表达分析及蛋白相互作用对其进行鉴定;通过抗生素胁迫、氧胁迫、生物被膜形成等实验研究Phd-Doc在体内发挥的生物学功能。【结果】生物信息学分析鉴定出一对Phd-Doc毒素-抗毒素系统,发现二者共转录且相互作用;毒素蛋白Doc能够引起大肠杆菌形态发生变化并抑制其生长,抗毒素蛋白Phd能中和Doc的毒性;Phd-Doc毒素-抗毒素系统具有自调控抑制效应;phd-doc的缺失对Yptb自身的生长无影响,而且毒素蛋白Doc在野生型Yptb内过表达并未显示毒性;phd-doc在转录水平上响应了抗生素胁迫和氧胁迫,其中,对氯霉素胁迫最为敏感,但并不影响Yptb的生长;同时,Phd-Doc能够影响Yptb的生物被膜形成能力。【结论】Yptb中Phd-Doc毒素-抗毒素系统的功能鉴定对于更好地了解在多变的外部环境下微生物的定殖和响应机制具有重要意义。     

Organ-specific distribution of dietary alkaloids in the marine opisthobranch Tylodina perversa

Specimens of the opisthobranch Tylodina perversa that were observed while feeding on the sponge Aplysina aerophoba were transferred to seawater tanks along with their prey and kept under controlled conditions. After one week the opisthobranchs were anaesthetized, dissected and studied for sequestered sponge-derived brominated alkaloids. All parts of T. perversa analyzed including feces, mucus and egg masses that had been produced during captivity contained alkaloids derived from A. aerophoba. The highest total alkaloid concentration (24.6 mg g⁻¹ dry wt) was found in mantles of T. perversa (compared to 51.2 mg g⁻¹ dry wt of total alkaloids in A. aerophoba). Hepatopancreas, egg masses and mucus (respective total alkaloid concentrations ranging from 20.4 to 12.5 mg g⁻¹ dry wt) were also rich in alkaloids. Whereas in A. aerophoba the isoxazoline alkaloids aerophobin-2 and isofistularin-3 were present in almost equal concentrations, aerophobin-2 constituted by far the major alkaloid (amounting to approximately 70% of all identified alkaloids) in mantles, mucus and egg masses of T. perversa, indicating selective sequestration by the opisthobranchs. Mantles as well as mucus also contained appreciable concentrations (approximately 20% of all identified compounds) of the brominated alkaloid aerothionin; this is not detected in A. aerophoba. It is possible that aerothionin originates from a previous encounter of T. perversa with the sponge A. cavernicola, the latter being closely related to A. aerophoba. The enrichment of aerophobin-2 (and of aerothionin) in mantles, mucus and egg masses that are vulnerable and exposed (mantles and egg masses) to predators and/or pathogens argues for defensive functions of the respective alkaloids even though this hypothesis still needs to be experimentally corroborated.     

Vitamin C stabilization as a consequence of the plasma membrane redox system

Summary Ascorbate is stabilized in the presence of HL-60 cells. Our results showed that cAMP derivatives and agents that increase cAMP stimulate the ability of HL-60 cells to stabilize ascorbate. On the other hand, tunicamycin, a glycosilation-interfering agent, inhibited this ability. The ascorbate stabilization in the presence of HL-60 cells has been questioned as a simple chemical effect. Further properties and controls about the enzymatic nature of this stabilization are described and discussed. This data, together with hormonal regulation, support the hypothesis that an enzymatic redox system located at the plasma membrane is responsible of the extracellular ascorbate stabilization by HL-60 cells.Abbreviations AFR ascorbate free radicals - FCS fetal calf serum - Sp-cAMPS Sp-cyclic adenosine monophosphothionate - Rp-cAMPS Rp-cyclic adenosine monophosphothionate     

Seasonal variations in the rates of aquatic and aerial respiration and ammonium excretion of the ribbed mussel, Geukensia demissa (Dillwyn)

Aquatic and aerial rates of oxygen consumption and ammonium excretion of ribbed mussels, Geukensia demissa (Dillwyn), collected from the mid-intertidal zone of a mid-Atlantic salt marsh, were measured under ambient conditions of food, temperature, and salinity over five seasons. Rates of aquatic respiration covaried with body size and season, as the rates were high and strongly related to mussel tissue weight in spring and summer but low and weight independent in winter. There was a significant interannual difference between summer of 1995 and 1996. Rates of aerial respiration also varied seasonally, with high rates of oxygen consumption in summer and low rates in winter. The magnitude of these seasonal variations were greater than those for aquatic respiration, and as a result, the ratio of aerial to aquatic respiration was higher in summer (0.61 and 0.52) than in winter (0.11). This indicates that G. demissa was able to actively regulate aerial respiration, thereby permitting high aerobic metabolism during prolonged periods of air exposure in summer. We hypothesize that such high rates of aerial respiration, during the seasons of high metabolic activity, are required to provide sufficient energy for mussels to facilitate food digestion during air exposure at low tide. Rates of ammonium excretion varied seasonally and increased with mussel weight in all seasons. The atomic ratio of oxygen to nitrogen (O:N), calculated from aquatic respiration vs. ammonium excretion, was significantly lower in autumn (26) than in other seasons (46–60) among which the O:N did not vary significantly.     

两种培养基对对虾苗池海洋蛭弧菌的分离及其多样性分析

【目的】明确海水(Sw)和聚蛋白胨20(Pp20)两种双层琼脂培养基对海洋蛭弧菌的分离计数效果,了解对虾苗池可培养海洋蛭弧菌多样性。【方法】采用双层平板法,比较Sw和Pp20培养基对2株海洋蛭弧菌和对虾苗池未知海洋蛭弧菌的计数效果。通过宿主范围测试和16S rRNA基因序列分析评估两种培养基分离苗池海洋蛭弧菌的多样性。【结果】宿主菌含量高时,Sw培养基对两株已知海洋蛭弧菌的计数值均显著高于(P0.05)Pp20。Sw和Pp20培养基从同一苗池水样分别分离得到21和22株蛭弧菌。根据宿主裂解范围差异,43株分离物可分为15种裂解模式,其中Sw和Pp20培养基各分离到12和8种。16S rRNA基因序列分析表明,所有分离物都被鉴定为噬菌弧菌属(Bacteriovorax)菌株,并可分为6个类群,Sw和Pp20培养基分别分离到6和4个类群。【结论】Sw培养基在分离计数海洋蛭弧菌及其多样性检测上效果均优于Pp20;对虾苗池可培养海洋蛭弧菌具有较高多样性,并以类群XIII、X及一个潜在新类群为优势种群。     

Study of selective feeding in the marine cladoceran Penilia avirostris by HPLC pigment analysis

Food selection by the marine cladoceran Penilia avirostris was studied in the field by HPLC analysis of phytoplankton marker pigments and in the laboratory by microscopic measurement of cell removal. Comparison between pigment composition in natural phytoplankton and in P. avirostris showed that P. avirostris preferred diatoms, cryptophytes and chlorophytes, and ignored prymnesiophytes and dinoflagellates. Peridinin, the marker pigment for dinoflagellates was found in P. avirostris only when the dinoflagellate populations were dominated by Prorocentrum. Pigment degradation rates ranged from 13.73% for alloxanthin to 36.62% for chlorophyll a. Clearance rates measured in the laboratory provided further evidence of strong preference for diatoms and cryptophytes, and avoidance of dinoflagellates. Microscopic counts suggested that P. avirostris was feeding on prymnesiophytes, although ingestion of prymnesiophytes could not be confirmed by pigment analysis.     

Oxidative stress in Perna perna and other bivalves as indicators of environmental stress in the Brazilian marine environment: Antioxidants, lipid peroxidation and DNA damage

Oxidative stress can take place in marine bivalves under a series of environmental adverse conditions. The study of different systems related to oxidative stress in these organisms can give important information about their physiological status and also about environmental health. Bivalves have been proposed as good sentinel organisms in pollution monitoring studies through the analysis of biochemical biomarkers, and most of the biomarkers analyzed are those related to oxidative stress. However, it is very important to know how other environmental factors not associated to the presence of pollutants might affect these parameters. We have studied a series of mechanisms related to oxidative stress in mussels which inhabit the Brazilian coast, especially in Perna perna species, subjected to different stress conditions, such as the exposure to different contaminants in the laboratory and in the field, the exposure of mussels to air and re-submersion, simulating the tidal oscillations, and in mussels collected at different seasons. Both oxidative damage levels and antioxidant defense systems were strongly affected by the different environmental stress. This review summarizes the data obtained in some studies carried out in bivalves from the Brazilian coast.     

The effect of sex hormones on lipid content and mast cell number in the harderian gland of the female toad, Bufo viridis

The Harderian gland of the toad Bufo viridis is a dimorphic gland owing to the presence of lipid droplets in the female glandular cells present only during summer months. Ovariectomy causes the disappearence of sudanophilia and estrogen-treatment completely prevents this change, while testosterone-injection has little effect. Estradiol-treatment also provokes a proliferation of the interstitial connective tissue concomitantly with the mast cell number increase. Our results suggest that estradiol acts, stimulating both mast cell and connective tissue proliferation, and plays a role in determining the expression of the female type of the toad Harderian gland.