and Its Expression in Pichia pastoris

利用RT-PCR技术扩增出草鱼(Ctenopharyngodon idellus)的leptin基因，将leptin基因克隆至真核表达载体pPIC9K，电穿孔转化GS115菌株, 经G418筛选和甲醇诱导后，对表达产物进行SDS-PAGE琼脂糖凝胶电泳和Western blot分析。结果表明，草鱼leptin基因cDNA序列由438个核苷酸组成，编码146个氨基酸组成的多肽（GenBank登陆号AY551335），与鲤鱼(Cyprinus carpio)leptin基因相比， 核苷酸和氨基酸的同源性为99%； 与人、猪和鼠相比，核苷酸同源性分别为84%、86%和95%, 氯基酸的同源性分别为84%、82%和96%；与河豚(Takifugu rubripes)相比，氨基酸具有较大的差异，仅有9％的同源性，表明leptin在物种的进化上具有一定的差异；实现了草鱼leptin基因在毕赤酵母(Pichia pastoris)中的表达，表达蛋白的分子量约为16 kD， Western blot 分析表明，表达产物具有一定的免疫学活性。     

Effects of Cd and Pb on the survival and juvenile production of Sinella coeca and Folsomia candida

Degradation factors can cause both quantitative and qualitative changes in fauna and thus affect soil functioning. The use of bio-indicators may help to detect environmental changes. The aim of this study was to evaluate the effects of lead and cadmium on the biology (mortality, reproduction, egg hatching) of two euedaphic collembolans, Sinella coeca and Folsomia candida and to evaluate sensitivity differences between the two species. The results showed that low concentrations of lead did not cause dramatic effects on the survival and reproduction of both species. Reproduction of both species was reduced when the soil contained a nominal concentration of Pb at 1000 μg g^–1. S. coeca and F. candida showed a high sensitivity to cadmium and both their reproduction and survival were significantly reduced on exposure to 50 μg g^–1. Moreover, egg hatching of F. candida was affected by the cadmium concentrations in the soil, and by the amount of time that the eggs stayed in contact with cadmium. This study showed that the inhibition determined by Cd on F. candida juvenile production was caused in part by inhibition of deposition and, in part, by reduction of egg viability.     

of Goat Follicle-stimulating Hormone Receptor (FSHR) Gene

利用PCR-SSCP技术检测山羊(Capra hircus)包括西农萨能奶山羊、关中奶山羊、陕南白山羊、安哥拉山羊和波尔山羊173个个体FSHR基因第10外显子的单核苷酸多态性 (SNP)。结果未发现SNP位点。测序后获得山羊FSHR基因第10外显子的核苷酸序列， 并在NCBI数据库中获得GenBank登录No.DQ069909和DQ069910。通过DNA序列分析发现， FSHR基因第10外显子第120位碱基不存在C→T的转换，也不存在颠换等其它遗传变化。山羊、绵羊 (Ovisaries L.) 和普通牛( Bos taurus) FSHR基因第10外显子序列同源性比较和聚类分析结果表明，山羊、普通牛和绵羊该部分序列的相似性最高为99.3%；在物种间比较中，绵羊和普通牛纯合子该基因外显子序列的不相似性最高为3.4%；据FSHR基因外显子序列构建的分子系统树结果显示，山羊、绵羊和普通牛物种内的个体各自聚为一类；山羊和绵羊先聚为一类，然后再与普通牛聚为一类。提示FSHR基因第10外显子的核苷酸序列适合于物种间的动物分子树的构建。     

The effect of two ant species Lasius niger and Lasius flavus on soil properties in two contrasting habitats

Ants significantly change the soil environment within the nest. The aim of this study is to contribute to ecology and thus the importance of two ant species Lasius niger and Lasius flavus in a post-mining landscape near the town of Sokolov in northwest Bohemia where both species are common. Chemical (total C, N, and available P) and microbiological parameters (respiration, cellulose decomposition and direct counts of bacteria) were investigated in both ant species in two different habitats: a tertiary clay heap after brown coal mining with a weakly developed organic layer and semi natural meadows with well developed organic horizons. Total C and N in the L. flavus mound was lower than in the surrounding soil in both stands, the same was true for total N in L. niger on the heaps. L. niger nests in both sites were significantly enriched by available P. A litter bag test with cellulose indicated lower decomposition in the ant nest in comparison with the surrounding soil. Respiration seems to be limited by lower soil moisture in the nest. However, microbial respiration, even in suitable moisture conditions, did not differ between the nest and soil (on heaps) or nest respiration was significantly lower (in L. flavus nests in the meadow). In meadow soil both species had a lower bacteria count than the surrounding soil, but the L. niger nest on the heap had higher bacterial numbers. Both species significantly alter soil conditions, although the effect on selected parameters is variable. Moreover, the result with lower nest moisture and lower decomposition rate in ant mounds indicates that soil moisture should be the next important factor limiting soil processes inside ant mounds.     

Effects of disturbance on the biodiversity and abundance of isopods in temperate grasslands

Connell's ‘intermediate disturbance hypothesis’ was tested using a relatively simple, species poor isopod community in grass heaths and in a coastal sand dune grassland. Variation in diversity resulted from disturbance both to the soil profile and sward structure due to grazing. Predictions from a spatial survey were tested using a 20 years time series. These changes were related to changes in rabbit activity, anthropogenic soil disturbance and on one site, deposition of blown sand. Changes in the soil profile resulted in diversity initially increasing due to decreases in the density of the commonest species, leading to an increase in equitability. Prolonged disturbance to the soil profile both due to sand blowing onto one site and rotavating on another, lead to a reduction in diversity due to one of the species being eliminated. The isopod community was most diverse when least disturbed by grazing, of intermediate diversity under intermediate grazing intensity and least diverse when most heavily grazed. In these simple communities, differences in diversity were mostly determined by differences in equitability rather than in species richness. We conclude that maximising heterogeneity of sward structure could have advantages for the conservation of both invertebrate biodiversity and vertebrates that feed on them.     

The influence of livestock dung on the abundance of exotic and native earthworms in a grassland in south-eastern Australia

It is well known that organic matter in the form of dung is utilised as a food source by some earthworms, but little has been reported on the preferences of earthworms for different types of dung in agricultural settings. An experiment was carried out in spring in south-eastern Australia to evaluate the effect that dung from different livestock has on the abundance of earthworms in a grassland environment. We were particularly interested to compare the responses of native Australian earthworms (Megascolecidae) with those of exotic earthworms (Lumbricidae and Acanthodrilidae). The attractiveness of dung from sheep, cattle and horses was measured by determining the abundance and biomass of the resident earthworm species under each dung type at varying times after adding the dung to the soil surface (0, 10, 20 and 30 days). The earthworm population consisted of three exotic species, Apporrectodea trapezoides, Microscolex dubius and M. phosphoreus, and two native species, Spenceriella macleayi and S. bywongensis. Both the number of days that the dung was available to earthworms and the type of dung influenced the numbers and biomass of the earthworms found beneath the dung pats. Significant interactions existed between time and dung type when all adult earthworms were considered as one group and also when adults were split into individual species. The various species responded differently to the dung, but horse dung was generally the more preferred dung type. The significance of these results is discussed in terms of the management of dung in an Australian pastoral context.     

Purification and isotopic signatures (<Emphasis Type="Italic">δ</Emphasis><Superscript>13</Superscript>C, <Emphasis Type="Italic">δ</Emphasis><Superscript>15</Superscript>N, Δ<Superscript>14</Superscript>C) of soil extracellular DNA

The aim of this work was to obtain pure extracellular DNA molecules so as to estimate their longevity in soil by an isotope-based approach. Extracellular DNA molecules were extracted from all horizons of a forest soil and purified by the procedure of Davis (Purification and precipitation of genomic DNA with phenol–chloroform and ethanol. In: Davis LG, Dibner MD, Battey JF (eds) Basic methods in molecular biology. Appleton & Lange, Norwalk, 16–22, 1986) without (DNA1) or with (DNA2) a successive treatment with binding resins followed by elution. The two differently purified DNA samples were compared for their A₂₆₀/A₂₈₀ ratio, polymerase chain reaction (PCR) amplification and natural abundance of stable (¹³C and ¹⁵N) and radioactive (¹⁴C) isotopes. The purity index and the PCR amplification did not differentiate the efficiency of the two purification procedures. The isotopic signature of DNA was more sensitive and was strongly affected by the purification procedures. The isotopic measurements showed that the major contaminant of extracellular DNA1 was the soil organic matter (SOM), even if it is not possible to exclude that the similar δ ¹³C, δ ¹⁵N and Δ¹⁴C values of DNA and SOM could be due to the use of SOM-deriving C and N atoms for the microbial synthesis of DNA. For extracellular DNA2, extremely low values of Δ¹⁴C were obtained, and this was ascribed to the presence of fossil fuel-derived substances used during the purification, although in amounts not revealed by gas chromatography-mass spectrometry analysis. The fact that it is not possible to obtain contaminant-free DNA molecules and the potential use of soil native organic compounds during the microbial synthesis of DNA make it not achievable to estimate the age of soil extracellular DNA by radiocarbon dating.     

Long-term effects of leguminous cover crops on biochemical and biological properties in the organic and mineral layers of soils of a coconut plantation

The primary aim of the study was to determine the long-term (12 years) effects of leguminous cover crops like Atylosia scarabaeoides, Centrosema pubescens, Calopogonium mucunoides and Pueraria phaseoloides on important soil biochemical and biological properties and their interrelationships in the organic (fresh litter layer, F and fermented + humus layer, F + H) and mineral (0–10 and 10–20 cm) layers of soils of a 19-year-old coconut plantation.The total biomass production (above-ground) for the 12-year period varied significantly between the cover crops and ranged from 34.86 (calopo) to 90.43 (pueraria) Mg ha^–1. Total N and C additions at the cover cropped (CC) site for the 12-year period were 0.97–3.07 Mg ha^–1 and 16.90–43.34 Mg ha^–1, respectively. Irrespective of layers, the levels of organic C, total N, organic substrates viz., dissolved organic C and N, labile organic N, water soluble carbohydrates, and light fraction organic matter-C and were markedly higher in the CC site compared to the control. Consequently, the levels of microbial biomass-C (C_MIC), -N (N_MIC) and -P (P_MIC), net N mineralization rates, CO₂ evolution, metabolic quotient (qCO₂) and the activities of l-asparaginase, l-glutaminase and β-glucosaminidase were significantly higher in the CC site compared to the corresponding levels in the control site. Between layers, the levels of various chemical, biochemical and microbial parameters were consistently higher in the organic layers compared to the mineral layers at all the sites including control. Among the ratios of various microbial indices, the ratios of C_MIC: organic C and C_MIC: P_MIC did not differ significantly between the layers and sites. However, the ratio of C_MIC: N_MIC was relatively higher in the mineral layers and control site. The variation in individual soil properties between layers and sites reflected the concomitant changes occurring in soil organic matter content. Apparently, microbial activity was limited by the supply of biologically available substrates in the mineral layers and the control site. Contrarily, the more direct supply of nutrients from decomposing plant litter and the indirect supply of nutrients from the mineralization of organic matter led to significantly higher levels of microbial biomass in the organic layers.     

Accumulation of heavy metals by enchytraeids and earthworms in a floodplain

The river floodplain ‘Afferdense and Deestsche Waarden’ (ADW) in The Netherlands is diffusely contaminated with several heavy metals. It is, however, unclear whether this mixed contamination exerts any adverse ecotoxicological effects. In November 2000 and May 2001 a field survey was conducted in two areas in the ADW to collect a wide range of data concerning contamination levels, bioavailability, enchytraeids and earthworms and abiotic factors such as lutum and organic matter content, cation exchange capacity (CEC) and soil nutrient concentrations. Earthworms and enchytraeids were also analyzed for heavy metal content. At both sites arsenic and zinc were present in soil at relatively high concentrations (above the Dutch intervention value). In the two areas, both enchytraeids and earthworms accumulated metals. Fridericia ulrikae accumulated more cadmium than Enchytraeus buchholzi and Henlea perpusilla. The earthworm Lumbricus rubellus accumulated larger concentrations of Cr, Cu and Pb than Aporrectodea caliginosa and Allolobophora chlorotica. Dietary, physiological and behavioral characteristics may have contributed to these differences.     

Production of Intergeneric Hybrids Between <Emphasis Type="Italic">Sinapis alba</Emphasis> and <Emphasis Type="Italic">Brassica carinata</Emphasis>

Reciprocal crosses were made between Brassica carinata and its related species Sinapis alba. Pollen germination studies indicated the presence post-fertilization barriers in both ways. Sequential ovary–ovule culture helped to realize the intergeneric hybrids from the cross S. alba × B. carinata. The F₁ obtained was confirmed as a hybrid based on morphology, cytology and isozyme studies. The hybrid was backcrossed to its male parent and obtained BC₁ seeds, which were used to raise BC₁ generation. The BC₁ generation plants were further backcrossed to B. carinata in order to develop alloplasmic lines.     

Climatic and litter fall effects on collembolan and oribatid mite species and communities in a beech wood based on a 7 years investigation

During a period of 7 years, between 1989 and 1995, Collembola and Oribatida were investigated in a beech forest on an acid Dystric Cambisol soil in northern Germany. Precipitation and temperature at a nearby climate station were recorded, and litter fall in the forest was measured. For 23 collembolan and 27 oribatid mite species, an analysis was performed concerning the influence of the climate parameters or litter fall on yearly assemblages or single species. Climate influence on the community structure was weak. Composition of the assemblages was relatively constant throughout the period of 7 years. A significant effect was determined for mean annual temperature and July precipitation only. More distinct effects were found in single species. In total, nine collembolan and six oribatid mite species were significantly influenced by litter fall, mean annual temperature, mean January temperature, mean July temperature, total precipitation or July precipitation. Reaction time ranged between 1 and 12 months. Four collembolan species reacted with a retarded yearly occurrence on deep spring temperature. Species diversity of Collembola was negatively correlated with total litter fall, while in oribatid mites the diversity showed a positive correlation.     

Nutrition on bacteria by bacterial-feeding nematodes and consequences on the structure of soil bacterial community

Bacterial-feeding nematodes are, with protozoa, the main grazers of soil bacteria. Interactions between bacteria and nematodes have important repercussions on soil functioning and particularly on nutrient availability. We assessed the influence of the bacterial strains ingested on bacterial-feeding nematodes population development and also the consequences of nematode feeding behaviour on the structure of the soil microbial community with a special attention to different soil micro-habitats for nematode and bacteria. In vivo studies conducted in the presence of single bacterial strains showed that the type of ingested bacteria conditioned the development of the different bacterial-feeding Cephalobidae nematode species tested and that the effect of bacteria differed between nematode species. The spatial distribution of soil nematodes between three soil habitats (fresh organic matter, inter-aggregates pores and aggregates) depended of the trophic behaviour of nematodes. Bacterial-feeding nematodes and fungal-feeding nematodes showed comparable distribution: being preferentially located in the fresh organic matter and in the inter-aggregate pores. Besides, the activity of inoculated bacterial-feeding nematodes modified the genetic structure of the soil microbial community. Bacterial community of the macroporosity was significantly influenced by the nematodes. On the contrary, no modification of the structure of the bacterial community linked with nematode activity was measured in the bulk soil.     

Response of CH<Subscript>4</Subscript> oxidation and methanotrophic diversity to NH<Subscript>4</Subscript><Superscript>+</Superscript> and CH<Subscript>4</Subscript> mixing ratios

Methane oxidising activity and community structure of 11, specifically targeted, methanotrophic species have been examined in an arable soil. Soils were sampled from three different field plots, receiving no fertilisation (C), compost (G) and mineral fertiliser (M), respectively. Incubation experiments were carried out with and without pre-incubation at elevated CH₄ mixing ratios (100 ml CH₄ l⁻¹) and with and without ammonium (100 mg N kg⁻¹) pre-incubation. Four months after fertilisation, plots C, G and M did not show significant differences in physicochemical properties and CH₄ oxidising activity. The total number of methanotrophs (determined as the sum the 11 specifically targeted methanotrophs) in the fresh soils was 17.0×10⁶, 13.7×10⁶ and 15.5×10⁶ cells g⁻¹ for treatment C, G and M, respectively. This corresponded to 0.11 to 0.32% of the total bacterial number. The CH₄ oxidising activity increased 10⁵-fold (20–26 mg CH₄ g⁻¹ h⁻¹), the total number of methanotrophs doubled (28–76×10⁶ cells g⁻¹) and the methanotrophic diversity markedly increased in treatments with a pre-incubation at elevated CH₄ concentrations. In all soils and treatments, type II methanotrophs (62–91%) outnumbered type I methanotrophs (9–38%). Methylocystis and Methylosinus species were always most abundant. After pre-incubation with ammonium, CH₄ oxidation was completely inhibited; however, no change in the methanotrophic community structure could be detected.     

Abundance of biogenic structures of earthworms and termites in a mango orchard

A comparative study of the spatial distributions and the quantity of biogenic structures produced by earthworms and termites (Odontotermes nilensis and Ancistrotermes guineensis) has been conducted in a mango orchard at Thiès (Senegal).This study showed that surface biogenic structures may represent a large amount of modified soil (up to 536.5 g m⁻²) which vary depending upon the seasons and the species. Whilst the quantity of casts was independent on the season (178.6 g m⁻²), O. nilensis sheetings fluctuated with the seasons. In addition, we show that the spatial organisation of surface biogenic structures fluctuates with seasons. It displays patches ranging from 5 to 15 m. There is a link between the distribution of earthworm casts and the vegetation. In addition, spatial distribution was also linked to the biology of constructing species. We observed that the A. guineensis’ filling structures were mainly located under the mango trees during the dry season where the stems and the brushwoods were abundant. It appears that the spatio-temporal distribution of the biogenic structures under study depended upon two main factors: season and vegetation. However, depending upon the biology of the engineer, these two factors influenced the spatial distribution of structures in different ways.     

Solar Decolorization of Methylene Blue by Magnetic MgFe<Subscript>2</Subscript>O<Subscript>4</Subscript>-MWCNT/Ag<Subscript>3</Subscript>VO<Subscript>4</Subscript> Visible Active Photocatalyst

MgFe₂O₄-MWCNT/Ag₃VO₄ photocatalyst was prepared for benefiting the visible region of solar spectrum. Prepared catalyst was characterized by using scanning electron microscope (SEM), X-ray diffraction (XRD), and energy-dispersive X-ray spectroscopy (EDX). Photocatalytic activity was measured by methylene blue (MB) decolorization under visible light obtained from a 105-W tungsten light bulb. Dye decolorization and its kinetics were followed up by means of a UV-vis spectrophotometer. Kinetic model of decolorization was found to be compatible with first-order kinetics. The effects of pH and concentration of MB solution on the decolorization efficiency were determined. Low and high pH conditions were found to be more effective in increasing the MB decolorization yield and rate. On the other hand, due to the low transparency of concentrated MB solutions, an increase on decolorization time and a lowering in decolorization yield were encountered. Thanks to the magnetic MgFe₂O₃ nanoparticles, 96% of the catalyst could be recovered by a simple magnetic bar. It was observed that simulated wastewater containing MB was also successfully decolorized showing that visible region-sensitive MgFe₂O₄-MWCNT/Ag₃VO₄ photocatalyst can be benefited as a potential, efficient, and reusable material for the removal organic pollutants in aquatic environment.     

Diversity among landraces of Indian snapmelon (<Emphasis Type="Italic">Cucumis melo</Emphasis> var. <Emphasis Type="Italic">momordica</Emphasis>)

Diversity among 36 snapmelon landraces, collected from 2 agro-ecological regions of India (9 agro-climatic sub-regions), was assayed using RAPD primers, morphological traits of plant habit and fruit, 2 yield-associated traits, pest and disease resistance and biochemical composition (TSS, ascorbic acid, titrable acidity). Typical differences among accessions were observed in plant and fruit characteristics and snapmelon germplasm with high titrable acidity and possessing resistance to downy mildew, Cucumber mosaic virus, Zucchini yellow mosaic virus, Papaya ringspot virus, Aphis gossypii and Meloidogyne incognita was noticed in the collection. RAPD based grouping analysis revealed that Indian snapmelon was rich in genetic variation and region and sub-region approach should be followed across India for acquisition of additional melon landraces. Accessions of var. agrestis and var. momordica clustered together and there was a separate cluster of the accessions of var. reticulatus. Comparative analysis of the genetic variability among Indian snapmelons and an array of previously characterized reference accessions of melon from Spain, Israel, Korea, Japan, Maldives, Iraq, Pakistan and India using SSRs showed that Indian snapmelon germplasm contained a high degree of unique genetic variability which was needed to be preserved to broaden the genetic base of melon germplasm available with the scientific community. N. P. S. Dhillon and Ranjana contributed equally to this work and are considered the first authors.     

Characterization of intergeneric hybrids of <Emphasis Type="Italic">Erianthus rockii</Emphasis> and <Emphasis Type="Italic">Saccharum</Emphasis> using molecular markers

Erianthus rockii, a wild relative of sugarcane, is drought and cold tolerant, and both are potentially important agronomic traits to the sugarcane industry worldwide. As such it is of interest as a source of parental germplasm to sugarcane breeders and is currently being used in sugarcane introgression programs in both China and Australia. To date morphological characters have been used to verify the putative hybrids produced. Two crosses were generated between two different Saccharum species and two E. rockii accessions. Over 400 AFLP markers were used to identify the intergeneric hybrid progeny as well as determine hybrid diversity. Both crosses generated hybrids but efficiency levels were very different and are probably related to the different Saccharum parent used in each cross. Cross 1 was between a Saccharum officinarum and E. rockii and generated 100% hybrid progeny. Cross 2, however, was between a sugarcane hybrid (S. officinarum × Saccharum spontaneum) and E. rockii and only 10% of the progeny were intergeneric hybrids. Inheritance of markers in the progeny was analysed and for both crosses there were equal numbers of markers from both parents indicating n + n transmission of chromosomes. This is the first verification of E. rockii hybrids with molecular markers. It may now be possible to exploit genes of value from E. rockii in sugarcane breeding programs.