Molecular characterization and developmental expression patterns of thyroid hormone receptors (TRs) and their responsiveness to TR agonist and antagonist in Rana nigromaculata

Considering some advantages of Rana nigromaculata as an experimental species, we propose that this species, like Xenopus laevis, could be used to assay thyroid hormone（TH） signaling disrupting actions. To validate the utilizability of R. nigromaculata, we investigated the responsiveness of R. nigromaculata to a TH receptor（TR） agonist（T3） and antagonist（amiodarone） by analyzing expression, based on characterizing TR cDNA and developmental expression patterns. With high levels of identity with the corresponding genes in X. laevis, both TRα and TRβ in R. nigromaculata exhibited roughly similar developmental expression patterns to those of X. laevis, in spite of some species-specific differences. Both TRα and TRβ expression had greater changes in the liver and intestine than in the tail and brain during metamorphosis. T3 exposure for 2 days induced more dramatic increases of TRβ expression in stage 27 than in stage34 tadpoles but not in stage 42 tadpoles, showing that the responsiveness of R. nigromaculata to TH decreased with development and disappeared at the onset of metamorphic climax.Corresponding to greater changes of TRβ expression in the liver and intestine than in the tail and brain during metamorphosis, the liver and intestine had higher responsiveness to exogenous T3 than the tail and brain. Amiodarone inhibited T3-induced TRβ expression. Our results show that R. nigromaculata can be used as a model species for assaying TH signaling disrupting actions by analyzing TRβ expression, and intestine tissues at stage 27 are ideal test materials due to high responsiveness and easy accessibility.     

稀有鮈鲫HMGR基因全长克隆及雄鱼经五氯酚暴露基因表达的分析

3-羟基-3-甲基戊二酰辅酶A还原酶(3-hydroxy-3-methylglutaryl coenzyme A reductase,HMGR)是甲羟戊酸(mevalonic acid,MVA)途径中的第一个关键限速酶.克隆稀有鮈鲫HMGR基因全长,分析该基因在不同组织及雄鱼经不同浓度五氯酚(pentachlorophenol,PCP)暴露的表达差异,探索PCP对类固醇激素前体(胆固醇)合成水平基因转录调控的内分泌干扰机制.采用同源克隆策略及cDNA末端快速扩增技术(rapid-amplification of cDNA ends,RACE),从稀有鮈鲫肝脏中首次克隆得到3 101碱基(bp)的HMGR基因cDNA全长,基因命名为GrHMGR(GenBank accession No:KF885724).GrHMGR编码884个氨基酸,系统发育树分析表明,GrHMGR编码的蛋白与其他鱼类来源的HMGR氨基酸序列同源性较高.Real-time PCR分析表明GrHMGR的表达有组织特异性,在大脑,性腺和肝脏中均有表达,性腺中表达量最高.稀有鮈鲫雄鱼经不同浓度PCP暴露后,GrHMGR在大脑和性腺的表达随PCP浓度的增加显著下降,然而在肝脏组织中表达趋势有所不同.HMGR的表达下降会减少胆固醇的合成.说明PCP暴露会通过干扰稀有鮈鲫的胆固醇合成途径,进而对内分泌系统产生影响.     

A putative octopamine/tyramine receptor mediating appetite in a hungry fly

In the blowfly Phormia regina, experience of simultaneous feeding with d-limonene exposure inhibits proboscis extension reflex (PER) due to decreased tyramine (TA) titer in the brain. To elucidate the molecular mechanism of TA signaling pathway related to the associated feeding behavior, we cloned cDNA encoding the octopamine/TA receptor (PregOAR/TAR). The deduced protein is composed of 607 amino acid residues and has 7 predicted transmembrane domains. Based on homology and phylogenetic analyses, this protein belongs to the OAR/TAR family. The PregOAR/TAR was mainly expressed in head, with low levels of expression in other tissues at adult stages. Gene expression profile is in agreement with a plethora of functions ascribed to TA in various insect tissues. The immunolabeled cell bodies and processes were localized in the medial protocerebrum, outer layer of lobula, antennal lobe, and subesophageal ganglion. These results suggest that decrease of TA level in the brain likely affects neurons expressing PregOAR/TAR, causing mediation of the sensitivity in the sensillum and/or output of motor neurons for PER.     

Development and growth in synanthropic species: plasticity and constraints

Urbanization poses serious extinction risks, yet some species thrive in urban environments. This may be due to a pronounced developmental plasticity in these taxa, since phenotypically, plastic organisms may better adjust to unpredictable urban food resources. We studied phenotypic plasticity in Nuctenea umbratica, a common European forest and urban vegetation spider. We subjected spiderlings to low (LF), medium (MF) and high (HF) food treatments and documented their growth and developmental trajectories into adulthood. Spiders from the three treatments had comparable numbers of instars and growth ratios, but differed in developmental periods. Longest developing LF spiders (♀?=?390, ♂?=?320 days) had the smallest adults, but MF (♀?=?300, ♂?=?240 days) and HF (♀?=?240, ♂?=?210 days) spiders reached comparable adult sizes through shorter development. While males and females had comparable instar numbers, females had longer development, higher growth ratios, adult sizes and mass; and while males adjusted their moulting to food availability, female moulting depended on specific mass, not food treatment. We discussed the patterns of Nuctenea sex-specific development and compared our results with published data on two other Holarctic urban colonizers (Larinioides sclopetarius, Zygiella x-notata) exhibiting high plasticity and fast generation turn-over. We conclude that despite relatively unconstrained developmental time in the laboratory enabling Nuctenea to achieve maximal mass and size—main female fitness proxies—their relatively fixed growth ratio and long generation turn-over may explain their lower success in urban environments.     

四环素抗生素对污泥中四环素抗性基因丰度和表达水平的作用影响

为研究四环素(TC)抗生素对其抗性基因(TC-ARGs)转录表达的作用影响,以从活性污泥中筛选获得的四环素抗性细菌(TRB)作为研究对象,采用荧光定量PCR和逆转录PCR方法检测了7种TC-ARGs,包括tet A、tet C、tet G、tet M、tet O、tet W和tet X基因的丰度和表达水平,并探讨了TC与TC-ARGs丰度及其表达水平之间的相关关系.结果表明,在整个培养周期内,tet A、tet G和tet W基因丰度随TC暴露浓度的增大总体呈现上升趋势,但其余TC-ARGs基因丰度整体波动较大. TC胁迫对不同TC-ARGs的转录表达水平作用影响差别较大,其中,tet A基因表达水平相对稳定,且随TC浓度的升高而上调,在TC浓度为100 mg·L~(-1)时,其上调倍数高达5. 3倍.在短期(1 d) TC胁迫下,TC-ARGs转录表达水平随TC浓度的升高整体呈现上调趋势.由相关性分析可知,tet A和tet W基因丰度与其转录表达水平之间存在显著的线性相关性,表明其基因丰度可在一定程度上衡量和评价其抗性表达水平,进而反映其功能活性和环境风险.     

The molecular diversity of arbuscular mycorrhizal fungi in the arsenic mining impacted sites in Hunan Province of China

Arbuscular mycorrhizal fungi(AMF) can establish a mutualistic association with most terrestrial plants even in heavy metal contaminated environments. It has been documented that high concentrations of toxic metals, such as arsenic(As) in soil could adversely affect the diversity and function of AMF. However, there are still gaps in understanding the community composition of AMF under long-term As contaminations. In the present study, six sampling sites with different As concentrations were selected in the Realgar mining area in Hunan Province of China. The AMF biodiversity in the rhizosphere soils of the dominant plant species was investigated by sequencing the nuclear small subunit ribosomal RNA(SSU rR NA) gene fragments using 454-pyrosequencing technique. A total of 11 AMF genera were identified,namely Rhizophagus, Glomus, Funneliformis, Acaulospora, Diversispora, Claroideoglomus, Scutellopora,Gigaspora, Ambispora, Praglomus, and Archaeospora, among which Glomus, Rhizophagus, and Claroideoglomus clarodeum were detected in all sampling sites, and Glomus was the dominant AMF genus in the Realgar mining area. Redundancy analysis indicated that soil pH, total As and Cd concentrations were the main factors influencing AMF community structure. There was a negative correlation between the AMF species richness and the total As concentration in the soil,but no significant correlation between the Shannon–Wiener index of the AMF and plants. Our study showed that high As concentrations can exert a selective effect on the AMF populations.     

Vulnerability and behavioral response to ultraviolet radiation in the components of a foliar mite prey–predator system

Ambient ultraviolet-B (UVB) radiation impacts plant-dwelling arthropods including herbivorous and predatory mites. However, the effects of UVB on prey?Cpredator systems, such as that between the herbivorous spider mite and predatory phytoseiid mite, are poorly understood. A comparative study was conducted to determine the vulnerability and behavioral responses of these mites to ultraviolet (UV) radiation. First, we analyzed dose?Cresponse (cumulative irradiance-mortality) curves for the eggs of phytoseiid mites (Neoseiulus californicus, Neoseiulus womersleyi, and Phytoseiulus persimilis) and the spider mite (Tetranychus urticae) to UVB radiation from a UV lamp. This indicated that the phytoseiid mites were more vulnerable than the spider mite, although P. persimilis was slightly more tolerant than the other two phytoseiid mites. Second, we compared the avoidance behavior of adult female N. californicus and two spider mite species (T. urticae, a lower leaf surface user; Panonychus citri, an upper leaf surface user) in response to solar UV and visible light. N. californicus actively avoided both types of radiation, whereas P. citri showed only minimal avoidance behavior. T. urticae actively avoided UV as well as N. californicus but exhibited a slow response to visible light as well as P. citri. Such variation in vulnerability and avoidance behavior accounts for differences in the species adaptations to solar UVB radiation. This may be the primary factor determining habitat use among these mites on host plant leaves, subsequently affecting accessibility by predators and also intraguild competition.     

利用氧化亚氮还原酶基因 (nosZ) 评价人工湿地系统中的反硝化菌

应用人工潜流湿地净化微污染地表水,出水用于补给人工景观河.利用定量PCR测定了湿地植物根际土壤和景观河底泥中16S rDNA和nosZ的丰度,并采用PCR-DGGE技术考察了样品中含nosZ基因的群落结构及其相似性.定量PCR结果表明,潜流湿地及人工景观河16S rDNA、nosZ平均绝对丰度(以DNA计)分别为1.91E+07、1.26E+06和2.68E+07、8.37E+05 copies.ng-1,以干土计时分别为1.45E+11、9.31E+09和5.31E+11、1.45E+10 copies.g-1.景观河底泥中微生物总量和反硝化菌数量要高于湿地根际土壤,但是后者nosZ相对丰度(3.8%～10.1%)则明显高于前者(1.7%～4.1%).根际土壤和底泥样品聚类分析相似度低,根际土壤优势菌种大部分与红杆菌目(Rhodobacearales)、根瘤菌目(Rhizobiales)和伯克氏菌目(Burkholderiales)的细菌相似,而底泥的优势菌种均为不可培养的微生物.     

气态甲醛暴露对神经生长因子表达影响的体内实验

为了探讨气态甲醛对小鼠组织中NGF表达的诱导作用,以小鼠肺和脑作为实验材料,应用RT-PCR方法检测甲醛染毒后小鼠肺和脑中NGF-mRNA的表达量.结果显示,小鼠脑中NGF表达量随着染毒时间的增加而增加;而肺中NGF在甲醛染毒1d后就达到最大,但随着时间的增加NGF表达量反而下降;故NGF在小鼠这2种组织中呈现不同的表达效应.为了进一步研究甲醛、NGF和哮喘之间的联系,构建了大鼠的哮喘模型,应用RT-PCR方法检测哮喘模型鼠肺中NGF的表达趋势.结果显示,在大鼠哮喘模型中,低浓度甲醛组(1.0mg·m-3)NGF表达量显著上调(p<0.01),而高浓度甲醛组(3.0mg·m-3)NGF表达量反而下降,仅略高于空白对照组(p>0.05).     

Molecular evidence of facultative intraguild predation by Monochamus titillator larvae (Coleoptera: Cerambycidae) on members of the southern pine beetle guild

The southern pine bark beetle guild (SPBG) is arguably the most destructive group of forest insects in the southeastern USA. This guild contains five species of bark beetles (Coleoptera: Curculionidae: Scolytinae): Dendroctonus frontalis, Dendroctonus terebrans, Ips avulsus, Ips calligraphus, and Ips grandicollis. A diverse community of illicit receivers is attracted to pheromones emitted by the SPBG, including the woodborers Monochamus carolinensis and Monochamus titillator (Coleoptera: Cerambycidae). These woodborers have been traditionally classified as resource competitors; however, laboratory assays suggest that larval M. carolinensis may be facultative intraguild predators of SPBG larvae. This study used polymerase chain reaction (PCR)-based molecular gut content analyses to characterize subcortical interactions between M. titillator and members of the SPBG. The half-lives of SPBG DNA were estimated in the laboratory prior to examining these interactions in the field. A total of 271 field-collected M. titillator larvae were analyzed and 26 (9.6?%) tested positive for DNA of members of the SPBG. Of these larvae, 25 (96.2?%) tested positive for I. grandicollis and one (3.8?%) for I. calligraphus. Failure to detect D. terebrans and D. frontalis was likely due to their absence in the field. I. avulsus was present, but primers developed using adult tissues failed to amplify larval tissue. Results from this study support the hypothesis that larval Monochamus spp. are facultative intraguild predators of bark beetle larvae. Additionally, this study demonstrates the capabilities of PCR in elucidating the interactions of cryptic forest insects and provides a tool to better understand mechanisms driving southern pine beetle guild population fluctuations.     

Piriformospora indica confers cadmium tolerance in Nicotiana tabacum

Piriformospora indica, a root-colonizing endophytic fungus of Sebacinales, promotes plant growth and confers resistance against biotic and abiotic stresses. In order to confirm the influence of P. indica on growth, proline, malondialdehyde (MDA), chlorophyll, and cadmium (Cd) amounts in Nicotiana tabacum under Cd stress, hydroponics, pot and field trials were conducted. The results showed that P. indica can store Cd in plant roots and reduce leaf Cd content, reduce the concentration of MDA, and increase the proline and chlorophyll content and the activities of catalase, peroxidase, and superoxide dismutase under hydroponic Cd stress. RT-PCR analysis showed that the relative expression level of genes Gsh2, TaPCS1, oas1, GPX, and Hsp70 in colonized plants was 4.3, 1.4, 2.9, 1.7, and 6.9 fold higher than in un-colonized plants respectively. Cd exposure significantly reduced un-colonized plants'' agronomic traits compared to P. indica-colonized ones. Our results suggested that P. indica can sequester Cd in roots, so that much less cadmium was transported to leaves, and the increased concentrations of antioxidant enzymes, pigments and proline contents, as well as the higher expression of stress-related phytochelatin biosynthesis genes in P. indica-inoculated plants, may also serve to protect N. tabacum plants against oxidative damage, enhancing Cd tolerance.     

Growth,morphology and yield of wheat,barley and oats grown at elevated atmospheric CO2 concentration in a cool,maritime climate

The effects of elevated CO₂ concentration on the growth, yield and quality of spring wheat (Triticum aestivum L., cv. ‘Sport’), barley (Hordeum vulgare, cv. ‘Thule’) and oats (Avena sativa, cv. ‘Kapp’) were studied. The study was performed from 20 April to 24 August in ten field chamber units each of 9 m² in a cool (12.6°C) maritime climate under long days (14.6–18.1 h), on the southwest coast of Norway (59°N, 6°E). The total biomass increased at high CO₂ concentration, by 11% and 20% in wheat and barley, respectively. The proportion of small grains increased by 6% in wheat and 26% in barley, but the total grain yield was not affected. The weight of chaff increased by 9% and 19% in wheat and barley, respectively. Plant height was significantly reduced during the growing season at elevated CO₂, by 8–19% in barley and by 9–25% in oats until 6 July when no significant difference in height was found. After 6 July, barley plants at elevated CO₂ were significantly taller than at ambient CO₂ concentration and oats were not affected. Elongation in wheat was not affected by CO₂ concentration at any time in the growing season. No difference in developmental rate could be detected between plants at normal and elevated CO₂ concentrations. The protein content of the grain decreased by 8% in barley, but was not significantly affected in the other species.     

Species-specific and female host-biased ectophoresy in the roundworm Caenorhabditis japonica

Caenorhabditis japonica is a bacteriophagous nematode species that was discovered on the semi-social burrower bug, Parastrachia japonensis, which demonstrates egg-guarding and provisioning behaviors. To understand the life history of C. japonica in relation to P. japonensis, we demonstrated the specificity of this association and fluctuations in nematode number on the insect throughout the year. C. japonica dauer larvae (DL), larvae in a nonfeeding diapause stage, were predominantly found as clumps on the adult female insects but rarely found on the male insects in all populations examined. This female-biased association was consistent throughout the year, but after the nymphs hatched, nematodes were not detected on the mother insects showing provisioning behavior. DL appeared on the nymphs, and the number of DL on the newly emerged female insects gradually increased thereafter. C. japonica has never been detected on other invertebrates collected from the P. japonensis habitat thus far. Our data suggest that the life cycles of C. japonica and P. japonensis are synchronized.     

四环素胁迫对Shigella flexneri细菌四环素抗性基因抗性表达的影响过程

四环素(TC)抗生素在不同环境介质中已被广泛检出,为研究其对四环素抗性基因(TC-ARGs)丰度变化及表达水平的影响过程,以从活性污泥中筛选和纯化分离获得的弗氏志贺氏菌(Shigella flexneri)为研究对象,考察了不同浓度TC对其生长过程的作用影响,采用荧光定量PCR和逆转录PCR方法定量检测了不同抗性机制TC-ARGs,包括tetC、tetO和tetX基因的丰度变化及表达水平,并探讨了TC浓度与TC-ARGs丰度及其表达水平之间的相关关系.结果表明,在培养周期内(24 h),TC胁迫对Shigella flexneri细菌的生长具有抑制作用,细菌细胞浓度增长速率随TC暴露浓度的升高而降低,但对TC-ARGs丰度变化影响较小.TC胁迫能够促进Shigella flexneri细菌TC-ARGs的转录表达,tetC、tetO和tetX基因表达水平在整个培养周期内均先升高后降低.由相关性分析可知,TC浓度与TC-ARGs丰度及其表达水平之间相关关系不显著,但tetC和tetO基因丰度与其转录表达水平之间存在显著的正相关关系,表明其基因丰度一定程度上可用来衡量和评价其抗性表达水平.     

Functional relationship between ammonia-oxidizing bacteria and ammonia-oxidizing archaea populations in the secondary treatment system of a full-scale municipal wastewater treatment plant

The abundance of ammonia-oxidizing bacteria and archaea and their amoA genes from the aerobic activated sludge tanks, recycled sludge and anaerobic digesters of a full-scale wastewater treatment plant (WWTP) was determined. Polymerase chain reaction and denaturing gradient gel electrophoresis were used to generate diversity profiles, which showed that each population had a consistent profile although the abundance of individual members varied. In the aerobic tanks, the ammonia-oxidizing bacterial (AOB) population was more than 350 times more abundant than the ammonia-oxidizing archaeal (AOA) population, however in the digesters, the AOA population was more than 10 times more abundant. Measuring the activity of the amoA gene expression of the two populations using RT-PCR also showed that the AOA amoA gene was more active in the digesters than in the activated sludge tanks. Using batch reactors and ddPCR, amoA activity could be measured and it was found that when the AOB amoA activity was inhibited in the anoxic reactors, the expression of the AOA amoA gene increased fourfold. This suggests that these two populations may have a cooperative relationship for the oxidation of ammonia.     

Toxic effects of perfluorononanoic acid on the development of Zebrafish (Danio rerio) embryos

Perfluorononanoic acid (PFNA) is a nine-carbon perfluoroalkyl acid widely used in industrial and domestic products. It is a persistent organic pollutant found in the environment as well as in the tissues of humans and wildlife. There is a concern that this chemical might be a developmental toxicant and teratogen in various ecosystems. In the present study, the toxic effects of PFNA were evaluated in zebrafish (Danio rerio) embryos. One hour post-fertilization embryos were treated with 0, 25, 50, 100, 200, 300, 350, and 400 μmol/L PFNA for 96 hr in 6-well plates. Developmental phenotypes and hatching rates were observed and recorded. Nineteen genes related to oxidative stress and lipid metabolism were examined using Quantitative RT-PCR and confirmed by whole mount in situ hybridization (WISH). Results showed that PFNA delayed the development of zebrafish embryos, reduced the hatching rate, and caused ventricular edema and malformation of the spine. In addition, the amount of reactive oxygen species in the embryo bodies increased significantly after exposure to PFNA compared with that of the control group. The Quantitative RT-PCR and WISH experiments demonstrated that mRNA expression of the lfabp and ucp2 genes increased significantly while that of sod1 and mt-nd1 decreased significantly after PFNA exposure. The mRNA expression levels of gpx1 and mt-atp6 decreased significantly in the high concentration group. However, the mRNA expression levels of both ppara and pparg did not show any significant variation after exposure. These findings suggest that PFNA affected the development of zebrafish embryos at relatively low concentrations.