Allelopathic inhibition of epiphytes by submerged macrophytes

The hypothesis that epiphytes are more vulnerable to allelochemicals released by submerged macrophytes than phytoplankton was tested by measuring growth and photosystem (PS) II activity of three common epiphytic algae and cyanobacteria in coexistence with Myriophyllum spicatum using dialysis tubes. Results were compared with earlier experiments on planktonic species. Contrary to the planktonic species, the tested epiphytes, the green algae Stigeoclonium tenue, the diatom Gomphonema parvulum and the cyanobacterium Oscillatoria limosa, were not significantly inhibited by M. spicatum. Growth and PS II activity of O. limosa were even significantly enhanced by M. spicatum, but this effect disappeared under phosphorus-deficiency due to the allelopathically induced inhibition of the alkaline phosphatase activity or phosphorus leakage by the macrophytes. My findings of a lower vulnerability of epiphytes against allelopathic substances of submerged macrophytes are supported by results of a literature survey.     

Rapid in vitro propagation,conservation and analysis of genetic stability of Viola pilosa

A protocol for in vitro propagation was developed for Viola pilosa, a plant of immense medicinal value. To start with in vitro propagation, the sterilized explants (buds) were cultured on MS basal medium supplemented with various concentrations of growth regulators. One of the medium compositions MS basal + 0.5 mg/l BA + 0.5 mg/l TDZ + 0.5 mg/l GA₃ gave best results for in vitro shoot bud establishment. Although the problem of shoot vitrification occurred on this medium but this was overcome by transferring the vitrified shoots on MS medium supplemented with 1 mg/l BA and 0.25 mg/l Kn. The same medium was found to be the best medium for further in vitro shoot multiplication. 100 % root induction from in vitro grown shoots was obtained on half strength MS medium supplemented with 1 mg/l IBA. In vitro formed plantlets were hardened and transferred to soil with 83 % survival. Additionally, conservation of in vitro multiplying shoots was also attempted using two different approaches namely slowing down the growth at low temperature and cryopreservation following vitrification. At low temperature retrieval rate was better at 10 °C than at 4 °C after conservation of in vitro multiplying shoots. In cryopreservation–vitrification studies, the vitrified shoot buds gave maximum retrieval of 41.66 % when they were precooled at 4 °C, while only 16.66 % vitrified shoots were retrieved from those precooled at 10 °C. Genetic stability of the in vitro grown plants was analysed by RAPD and ISSR markers which indicated no somaclonal variation among in vitro grown plants demonstrating the feasibility of using the protocol without any adverse genetical effects.     

Environmental factors regulating the radial oxygen loss from roots of Myriophyllum spicatum and Potamogeton crispus

Myriophyllum spicatum and Potamogeton crispus are common species of shallow eutrophic lakes in north-eastern Germany, where a slow recovery of the submersed aquatic vegetation was observed. Thus, the characterisation of the root oxygen release (ROL) as well as its implication for geochemical processes in the sediment are of particular interest. A combination of microelectrode measurements, methylene blue agar and a titanium(III) redox buffer was used to investigate the influence of the oxygen content in the water column on ROL, diel ROL dynamics as well as the impact of sediment milieu. Oxygen gradients around the roots revealed a maximum oxygen diffusion zone of up to 250 μm. During a sequence with a light/dark cycle as well as alternating aeration of the water column, maximum ROL with up to 35% oxygen saturation at the root surface occurred under light/O₂-saturated conditions. A decrease to about 30% was observed under dark/O₂-saturated conditions, no ROL was detected at dark/O₂-depleted conditions and only a weak ROL with 5–10% oxygen saturation at the root surface was measured under light but O₂-depleted water column. These results indicate, that during darkness, ROL is supplied by oxygen from the water column and even during illumination and active photosynthesis production, ROL is modified by the oxygen content in the water column. Visualisation of ROL patterns revealed an enhanced ROL for plants which were grown in sulfidic littoral sediment in comparison to plants grown in pure quartz sand. For both plant species grown in sulfidic littoral sediment, a ROL rate of 3–4 μmol O₂ h⁻¹ plant⁻¹ was determined with the Ti(III) redox buffer. For plants grown in pure quartz sand, the ROL rate decreased to 1–2 μmol O₂ h⁻¹ plant⁻¹. Hence, aside from the oxygen content in the water column, the redox conditions and microbial oxygen demand in the sediment has to be considered as a further major determinant of ROL.     

The effect of temperature, water level and burial depth on seed germination of Myriophyllum spicatum and Potamogeton malaianus

The effects of temperature, water level and burial depth on seed germination of two submerged species, Myriophyllum spicatum and Potamogeton malaianus, were investigated under controlled laboratory conditions. There was no significant difference in final germination of M. spicatum among water level treatments, but P. malaianus germinations at 1 cm and 12 cm water levels were better than at 0 cm water level at temperatures of 20 °C and 30 °C. Little to no germination was observed for either species at the temperature of 10 °C. At 15 °C, however, germination increased significantly to 66.3-70.6% for M. spicatum and to 29.4-48.1% for P. malaianus under all three water level treatments. Increased temperature from 15 °C to 30 °C had no significant effect on the final germination of M. spicatum except at the 1 cm water level, but enhanced significantly the germination of P. malaianus. Analysis of the mean time to germination revealed that M. spicatum was a faster germinator relative to P. malaianus. The two species’ germination differed markedly in response to burial depth. Germination percentage of M. spicatum was 71.3% at 0 cm burial depth, but decreased to 5.0% and to 2.5% at depths of 1 cm and 2 cm, respectively; whereas germination percentages of P. malaianus were 40.0%, 23.8%, 12.5%, 7.5% and 1.3% at depths of 0 cm, 1 cm, 2 cm, 3 cm and 5 cm, respectively. We concluded that the two species respond differently to germination strategies. The findings provided further insight into how germination strategy contributes to the seed bank formation and species invasion.     

Growth and nutrient absorption of two submerged aquatic macrophytes in mesocosms, for reinsertion in a eutrophicated shallow lake

Aquatic macrophytes play a central role in preserving the ecological equilibrium of shallow lakes and in the restoration of eutrophic lakes that have switched to phytoplankton-dominated turbid water. Massaciuccoli Lake, a shallow lake located along the Tuscan coast in Italy, has shown a constant and progressive simplification of the submerged plant community, for anthropogenic reasons, leading, in recent years, to turbid water. The growth and nutrient absorption capability of two macrophyte species, Myriophyllum verticillatum L. and Elodea canadensis Michaux, in the lake was investigated, with the prospect of a future lake restoration programme centred on their replacement. Mesocosm experiments were conducted to monitor the plant growth and nutrient (NO₂⁻, NO₃⁻, NH₄⁺, Ntot, PO₄³⁻, Ptot) content in the plant dry matter and water at the beginning and at the end of the trial. Bacterial activity was analysed in the water in order to verify the possible nutrient absorption contribution by organisms other than plants. Both M. verticillatum and E. canadensis showed satisfactory growth and nutrient reduction in the water body. Moreover, their different growth patterns suggested that optimal replacement can be performed with their introduction in two steps, starting with M. verticillatum, which shows the best capacity to colonise the aquatic environment, due to its tendency towards lengthening.     

太湖蓝藻死亡腐烂产物对狐尾藻和水质的影响

为揭示富营养化造成淡水生态系统中沉水植物消亡的机制,本文就太湖蓝藻大规模死亡腐烂后的产物对水体水质以及沉水植物造成的影响进行了研究。本文以沉水植物穗花狐尾藻为研究对象,测定了水质参数（pH值,浊度（NTU）,温度,溶解氧（DO）,电导率（Ec）,盐度,磷酸根（PO43-））以及狐尾藻Fv/Fm指标。将采自太湖梅梁湾的蓝藻水华,降解一个星期左右。随后在处理组中加入该蓝藻降解物,对照组中不加入。结果表明,加入蓝藻死亡腐解液后,水体的溶解氧、pH值均比对照组显著降低,浊度则显著升高。对照组中穗花狐尾藻生长良好,其Fv/Fm约为0.8,而经蓝藻腐烂液处理的实验组,穗花狐尾藻在两天之内便接近死亡状态,其Fv/Fm值降至0.3左右。以上结果表明蓝藻死亡腐烂后形成的腐解液确实能在很短时间内给沉水植被带来灭顶之灾。而在短时间内并不能形成大量附着生物,故本实验可排除其对水生植物的影响。本文证实了蓝藻死亡腐烂后的降解液极有可能是导致沉水植物消亡的主要原因。至于是由于蓝藻腐解液引起的哪种因素起主导作用,还有待进一步研究。     

The effects of floating mats of Azolla filiculoides Lam. and Lemna minuta Kunth on the growth of submerged macrophytes

In laboratory experiments, the growth characteristics of the submerged species Elodea nuttallii (Planch.) St. John and Potamogeton crispus L. were assessed in the presence and absence of floating mats of Azolla filiculoides Lam. and Lemna minuta Kunth. Light penetration and the development of pH and dissolved oxygen differences were monitored. The growth of P. crispus was suppressed much more than that of E. nuttallii and the effects of A. filiculoides were more severe than those of L. minuta. Findings are related to possible field responses of submerged plants under floating mats, especially their abilities to compensate for the potential suppressive effects of floating mats under natural conditions.     

Evolution of Cryptosporidium in vitro culture

This overview discusses findings from culturing Cryptosporidium spp. in cell and axenic cultures as well as factors limiting the development of this parasite in cultivation systems during recent years. A systematic review is undertaken of findings regarding the life cycle of the parasite, taking into account physiological, biochemical and genetic aspects, in the hope that this attempt will facilitate future approaches to research and developments in the understanding of Cryptosporidium biology.     

Development in vitro of the neuromusculature of two strigeid trematodes,Apatemon cobitidis proterorhini and Cotylurus erraticus

Confocal microscopy interfaced with cytochemical procedures has been used to monitor development of the major muscle systems and associated serotoninergic (5-HT, 5-hydroxytryptamine) and peptidergic (FaRP, FMRFamide-related peptide) innervation of the strigeid trematodes, Apatemon cobitidis proterorhini and Cotylurus erraticus during cultivation in vitro. Sexually undifferentiated metacercariae were successfully grown to ovigerous adults using tissue culture medium NCTC 135, chicken serum and egg albumen. Eggs were produced after 5 days in culture but had abnormal shells and failed to embryonate. 5-HT and FaRP (the flatworm FaRP, GYIRFamide) were localised immunocytochemically in both central and peripheral nervous systems of developing worms. During cultivation, the central serotoninergic and FaRPergic neuronal pathways of the forebody became more extensive, but retained the same basic orthogonal arrangement as found in the excysted metacercaria. Longitudinal extensor and flexor muscles of the hindbody provide support for the developing reproductive complex. The male reproductive tracts were established in advance (day 3) of those of the female system (day 4); completion of the latter was marked by the appearance of the ootype/egg chamber. The inner longitudinal muscle fibres of the female tract appeared prior to the outer and more densely arranged circular muscles. Circular fibres dominate the muscle complement of both alimentary and reproductive tracts. 5-HT- and GYIRFamide-immunoreactivities were demonstrable in the central nervous system (CNS) and subtegumental parasympathetic nervous system (PNS) throughout the culture period, but innervation of the developing reproductive structures was reactive just for 5-HT. Only at the onset of egg production was FaRP-IR observed in the reproductive system and was expressed only in the innervation of the ootype, a finding consistent with the view that FaRPs may regulate egg assembly in platyhelminths.     

Light-dependent phosphate uptake of a submersed macrophyte Myriophyllum spicatum L.

The uptake kinetics of phosphate (Pi) by Myriophyllum spicatum was determined from adsorption and absorption under light and dark conditions. Pi uptake was light dependent and showed saturation following the Michaelis-Menten relation (in light: V = 16.91 × [Pi](1.335 + [Pi]), R² = 0.90, p < 0.001; in the dark: V = 5.13 × [Pi](0.351 + [Pi]), R² = 0.77, p < 0.001). Around 77% of the loss of Pi in the water column was absorbed into the tissue of M. spicatum, and only 23% was adsorbed on the surface of the plant shoots. Our study shows that M. spicatum shoots have a much higher affinity (in light: 3.9 μmol g⁻¹ dw h⁻¹ μM⁻¹; in the dark: 3.7 μmol g⁻¹ dw h⁻¹ μM⁻¹) and V_max (maximum uptake rate, shoot light) for Pi uptake than many other aquatic macrophytes (in light: 0.002-0.23 μmol g⁻¹ dw h⁻¹ μM⁻¹; in the dark: 0.002-0.19 μmol g⁻¹ dw h⁻¹ μM⁻¹), which may provide a competitive advantage over other macrophytes across a wide range of Pi concentrations.     

In vitro production of huperzine A, a promising drug candidate for Alzheimer's disease

Alzheimer's disease (AD) is growing in impact on human health. With no known cure, AD is one of the most expensive diseases in the world to treat. Huperzine A (HupA), a anti-AD drug candidate from the traditional Chinese medicine Qian Ceng Ta (Huperzia serrata), has been shown to be a powerful and selective inhibitor of acetylcholinesterase and has attracted widespread attention because of its unique pharmacological activities and low toxicity. As a result, HupA is becoming an important lead compound for drugs to treat AD. HupA is obtained naturally from very limited and slowly growing natural resources, members of the Huperziaceae. Unfortunately, the content of HupA is very low in the raw plant material. This has led to strong interest in developing sources of HupA. We have developed a method to propagate in vitro tissues of Phlegmariurus squarrosus, a member of the Huperziaceae, that produce high levels of HupA. The in vitro propagated tissues produce even higher levels of HupA than the natural plant, and may represent an excellent source for HupA.     

Comparison of two methods characterising the seed bank of amphibious plants in submerged sediments

We analysed the submerged soil seed bank of three fishponds in the Waldviertel region in Austria. We aimed our study at comparing the efficiency of two methods in detecting seeds quantitatively from soil samples of four characteristic mud-flat species, i.e. Carex bohemica, Coleanthus subtilis, Elatine hexandra, and Eleocharis ovata: the seedling-emergence and the rinsing method. Additionally, the actual vegetation was described based on 65 phytosociological relevés. Soil seed bank analyses were based on 31 samples representing the vegetation zonation. Mixed soil samples were portioned into 62 subsamples further treated with the two methods in parallel.     

In vitro propagation of female Ephedra foliata Boiss. & Kotschy ex Boiss.: an endemic and threatened Gymnosperm of the Thar Desert

Ephedra foliata Boiss. & Kotschy ex Boiss., (family – Ephedraceae), is an ecologically and economically important threatened Gymnosperm of the Indian Thar Desert. A method for micropropagation of E. foliata using nodal explant of mature female plant has been developed. Maximum bud-break (90 %) of the explant was obtained on MS medium supplemented with 1.5 mg l⁻¹ of benzyl adenine (BA) + additives. Explant produces 5.3 ± 0.40 shoots from single node with 3.25 ± 0.29 cm length. The multiplication of shoots in culture was affected by salt composition of media, types and concentrations of plant growth regulators (PGR’s) and their interactions, time of transfer of the cultures. Maximum number of shoots (26.3 ± 0.82 per culture vessel) were regenerated on MS medium modified by reducing the concentration of nitrates to half supplemented with 200 mg l⁻¹ ammonium sulphate {(NH₄) ₂SO₄} (MMS3) + BA (0.25 mg l⁻¹), Kinetin (Kin; 0.25 mg l⁻¹), Indole-3-acetic acid (IAA; 0.1 mg l⁻¹) and additives. The in vitro produced shoots rooted under ex vitro on soilrite moistened with one-fourth strength of MS macro salts in screw cap bottles by treating the shoot base (s) with 500 mg l⁻¹ of Indole-3-butyric acid (IBA) for 5 min. The micropropagated plants were hardened in the green house. The described protocol can be applicable for (i) large scale plant production (ii) establishment of plants in natural habitat and (iii) germplasm conservation of this endemic Gymnosperm of arid regions.     

Chemical profile of the North American native Myriophyllum sibiricum compared to the invasive M. spicatum

Myriophyllum spicatum L. is a nonindigenous invasive plant in North America that can displace the closely related native Myriophyllum sibiricum Komarov. We analyzed the chemical composition (including: C, N, P, polyphenols, lignin, nonpolar extractables, and sugars) of M. spicatum and M. sibiricum and determined how the chemistry of the two species varied by plant part with growing environment (lake versus tank), irradiance (full sun versus 50% shading), and season (July through September). M. spicatum had higher concentrations of carbon, polyphenols and lignin (C: 47%; polyphenols: 5.5%; lignin: 18%) than M. sibiricum (C: 42%; polyphenols: 3.7%; lignin: 9%) while M. sibiricum had a higher concentration of ash under all conditions (12% versus 8% for M. spicatum). Apical meristems of both species had the highest concentration of carbon, polyphenols, and tellimagrandin II, followed by leaves and stems. Tellimagrandin II was present in apical meristems of both M. spicatum (24.6 mg g⁻¹ dm) and M. sibiricum (11.1 mg g⁻¹ dm). Variation in irradiance from 490 (shade) to 940 (sun) μmol of photons m⁻² s⁻¹ had no effect on C, N, and polyphenol concentrations, suggesting that light levels above 490 μmol of photons m⁻² s⁻¹ do not alter chemical composition. The higher concentration of polyphenols and lignin in M. spicatum relative to M. sibiricum may provide advantages that facilitate invasion and displacement of native plants.     

Physical control of Eurasian watermilfoil in an oligotrophic lake

The introduction of Eurasian watermilfoil (Myriophyllum spicatum) into oligotrophic waters of high water clarity in temperate zones of North America has produced growth in excess of 6 m depth and yearly biomass approaching 1000 g m^–2 dry weight. From its initial observation in Lake George, New York, USA in 1985, by 1993 milfoil had spread to 106 discrete locations within the lake. A 7-year study of one site having no management showed milfoil to grow expansively, suppressing native plant species from 20 in 1987 to 6 in 1993 with the average number of species m^–2 quadrat declining from 5.5 in 1987 to less than 2 in 1993. Management of milfoil by means of hand harvesting, suction harvesting and benthic barrier has reduced the number of unmanaged sites from 106 in 1993 to 11. One year post-treatment at sites utilizing suction harvesting, showed a greater number of native species at all sites than pretreatment with a substantial reduction in milfoil biomass. At sites where benthic barrier was removed 1–2 years after installation, milfoil had recolonized 44% of grid squares within 30 days. Ninety days after barrier removal 74% of grid squares contained milfoil and one year later 71% of the grids supported milfoil. During the first year following mat removal, the average number of species m^–2 peaked at 4.7 and stabilized at 4.5 during the second year. Hand harvesting by SCUBA in areas of limited milfoil growth (new sites of infestation and sites of former treatment) was found to reduce the number of milfoil plants present in subsequent years. Hand harvesting did not eliminate milfoil at any of the sites and regrowth/colonization necessitated reharvesting every 3 or more years. Results of evaluations of physical plant management techniques indicate that (1) an integrated program utilizing different techniques based on plant density reduced the growth of milfoil and (2) long term commitment to aquatic plant management is necessary since none of the techniques employed singly were found to eliminate milfoil.     

Use of Geographic Information Systems to monitor and predict non-native aquatic plant dispersal through north-eastern North America

North America has a growing problem with invasive aquatic plants. At every level of the aquatic food web, long-standing checks and balances have become dramatically eroded by the introduction of non-native species. The northeastern United States contains thousands of diverse freshwater habitats, highly heterogeneous in geology and locale, where nuisance aquatic plant growth results in decreased lake water quality, interference with recreational access, degraded flood control structures, and impacts to their aesthetic quality. Early infestation and spread of exotic species is often poorly documented at local and state levels. Consequently, successful management of these species depends on continuous monitoring and definitive identification by both public officials and waterside homeowners. With new mapping capabilities employing software such as Geographic Information Systems (GIS) coupled with the growing computerization of taxonomic records and online availability of regional herbarium records, we have been able to illustrate the␣temporal and geographic spread of these species, thus enabling aquatic ecologists and managers to make predictions of future infestations. In this paper we evaluate these techniques focusing on three of the most pervasive of exotic aquatic plant species in New York State: Myriophyllum spicatum, Trapa natans and Potamogeton crispus.     

In vitro propagation of green-foliaged Dracaena fragrans Ker.

Callus tissue was induced in young stem segments cultured on MS based media supplemented with 0.25–0.5 mg l^-1 2, 4-D. Shoots were differentiated on media containing 0.5–1.0 mg l^-1 BA and 0.5–2.0 mg l^-1 IBA or 0.1–0.2 mg l^-1 NAA. The same media were suitable for shoot multiplication. Shoot elongation and rooting were strongly inhibited by BA and stimulated by auxins IBA and NAA. Medium containing 0.5 mg l^-1 IBA was optimal for rooting. Root elongation was stimulated by light and inhibited in darkness. Transfer of rooted plantlets to outdoor conditions was feasible and special hardening procedures were not required. Among more than 5000 plants produced by this procedure only 9 off-type plants with variegated leaves were found.     

Promoting low-canopy macrophytes to compromise conservation and recreational navigation in a shallow lake

The shift from a turbid-water state to a clear macrophyte-dominatedstate in the shallow lake Veluwemeer (The Netherlands) has led to nuisance forrecreational navigation. The nuisance concerns the dense beds ofPotamogeton perfoliatus in particular, whereas thelow-canopy forming charophytes cause much less harm. On the other hand, theimportance of macrophyte cover for the stability of the clear-water state hasbeen recognised. To assess the potential of mechanical removal of densemacrophyte beds, several cutting regimes were simulated in a mixed vegetation ofP. perfoliatus and Chara aspera, usingthe individual-based model Charisma. These species occupy a wide range of waterdepths between 0.5 and 2.5 m, with C. asperadominating the shallower zone and P. perfoliatus dominatingthe deeper zone; intermediary is a zone where either species may dominate asalternative equilibria. Both the cutting height and timing affected the amountof biomass present in summer. The effect of cutting was more profound fortreatment later in the season. With a cutting level above theChara-canopy, the simulations showed an increased biomassof C. aspera and reduction of P.perfoliatus. In the zone of alternative equilibria, it was possibleto provoke a sustainable shift from P. perfoliatusdominance to C. aspera dominance. To achieve this, annualrepetition of cutting for a number of years was necessary. A harvesting regimeaimed at shifting the vegetation dominance from P.perfoliatus towards C. aspera could be an optionfor management of the lake, since it holds the perspective of decreasedmanagement effort after a number of years. Moreover, removing only high-canopyvegetation implies high conservation values and recreational use can becombined.     

菹草种群内外水质日变化

对菹草(Potamogeton crispus)种群内外水体进行了昼夜连续监测,分析菹草种群内外水质的日变化趋势.结果显示,种群内DO自日出后增加,日落后持续下降,且与水温变化一致,于19:30左右出现1次明显低谷,后略有恢复,后一直降至日出前后;开阔水域DO变化与水温呈现一定相关性,但变化幅度较小;交界处DO含量兼有种群内与开阔水域的变化特点.种群内水体pH值自日出前后升高,日落时达最高值,后开始下降,直至次日出前后;交界处与种群变化趋势一致;开阔水域pH总体变化幅度较小,白天高于夜间,总体DO含量及pH分布:种群区＞交界处＞开阔水域.种群及交界处TN含量均在日出前后达最高,日出后递减,开阔水域夜间含量较高,日出前后达最高;各点NH+4-N变化规律不显著;种群内水体TP含量夜间较高,日出前后达最高值,日出后递减,日落后递增;开阔水域TP含量夜间较高,白天略低;交界处正午前后达最低值,傍晚达最高值,TN、TP总体分布:种群＜交界处＜开阔水域.菹草种群存在对水体DO、pH等环境因子均产生重要影响,继而影响水体内源性氮磷的迁移,其中对pH影响较小,其变化未能影响水体氮磷迁移,而DO昼夜变化较大,对水体内源性氮磷的迁移起重要影响.     

Uptake of stable N isotopes by Myriophyllum spicatum is not selective

Preference for ¹⁴N over ¹⁵N was assessed for Myriophyllum spicatum L. by exposing shoots to the same initial concentration of N, but different isotopic compositions (100% ¹⁴N, 50% ¹⁴N and 50% ¹⁵N, and 100% ¹⁵N) for 6 weeks. Regardless of treatment, all available inorganic N was taken up by M. spicatum at similar rates. Significant differences in tissue isotopic composition were evident across treatments in 2 weeks, and persisted for the remainder of the experiment. At 6 weeks, atom (at) % ¹⁵N was twice as high in the 100% ¹⁵N treatment as in the 50% ¹⁵N treatment, and the at% of the ¹⁴N treatment (0% ¹⁵N) remained unchanged. This observed absence of preference for ¹⁴N in M. spicatum suggests that this species can be useful in determining N sources to freshwater systems, which often differ in ¹⁵N/¹⁴N ratio.