Grazing on bacteria by flagellates and cladocerans in lakes of contrasting food-web structure

We tested the hypothesis that grazing on bacteria would varybetween lakes with differing plankton community structures.Paul and Tuesday lakes (Gogebic County, MI) are respectivelydominated by piscivorous and planktivorous fish. Consequently,zooplankton in Paul are primarily large daphnids, while zooplanktonin Tuesday are primarily small cladocerans and copepods. Wemeasured flagellate grazing on bacteria using a fluorescentminicell method, while cladoceran grazing was estimated fromthe relationship between body length and filtering rate. Wepredicted that cladoceran grazing on bacteria would be higherin Paul, and flagellate grazing would be higher in Tuesday.Cladoceran grazing on bacteria was important in both lakes contraryto our initial expectation. Large populations of the small cladoceran,Bosmina longirostris, in Tuesday exerted a grazing pressure(0.18–35x10⁶ bacteria 1^–1 h^–1) approximatelyequal to that of the large cladoceran, Daphnia pulex, in Paul(0.34–30x10⁶ bacteria 1^–1 h^–1). Flagellategrazing was higher in Tuesday as predicted (range: Paul, 0.1–6x10⁶bacteria 1^–1 h^–1; Tuesday, 0.2–20x10⁶ bacteria1^–1 h^–1). However, there was not a simple relationshipbetween total abundance of flagellates and total grazing rates.High community grazing by flagellates occurred when attachedchoanoflagellates were present. These flagellates had higheringestion rates than free forms. We find no clear evidence thatdifferences in food-web structure between the two lakes influencethe process of grazing on bacteria. Instead, our results emphasizethe significance of cladocerans and attached flagellates asconsumers of bacteria in freshwater ecosystems.     

Growth and production of heterotrophic nanoflagellates in a meso-eutrophic lake

The growth of heterotrophic nanoflagellates (HNF) in mesotrophicLake Constance was measured in situ during a 13 month period.Experiments were conducted with 10 µm pre-filtered lakewater incubated in diffusion chambers at 3 m water depth atthe sampling location for 24 h. Growth rates were calculatedfrom changes in cell numbers occurring during the period ofincubation. Growth rates of all dominant taxa showed pronouncedseasonal variation (–0.13 to 1.76 day^–1 and weregenerally highest in summer at high water temperatures. In situgrowth rates were well below maximum growth rates known forthe respective and similar species from laboratory experiments.While water temperature was a key parameter positively relatedto the growth of all HNF species, the effect of various potentialfood items was taxon specific and less clear. Bacterial abundancewas equally important as temperature for growth in the smallbactenvorous Spumella sp., but was insignificant for growthrates of the larger omnivorous Kathablepharis sp. In Spuniellasp., 84% of the observed seasonal variation of its growth ratecould be explained by temperature and bacterial food supply.Based on these results, a multiple linear regression equationwith temperature and bacterial concentration as dependent variableswas calculated for the growth rate of Spumella. Taxon-specificproduction rates were derived from growth rates and averagebiomass of these two species, and compared to total HNF productionestimated from previously measured community growth rates andbiomass in Lake Constance. Production peaks of Spumella sp.and Kathablepharis sp. alternated seasonally. Total HINF productionranged from –0.01 to 10 mg C m^–3 day^–1. Theaverage seasonal production varied between 1.4 and 33 mg C m^–3day^–1 over 6 consecutive years. These small protozoa thuscontribute a substantial amount to total zooplankton productionin Lake Constance.     

Nitrogen Fixation, Assimilation and Transport During the Initial Growth Stage of Phaseolus vulgaris L

In nodulated common bean (Phaseolus vulgaris L.), there is typicallya period of N stress between 15 and 20 d after emergence (DAE),due to a lack of synchronization between the depletion of Nin the cotyledons and the beginning of N² fixation and transport.Screening trials identified some Rhizobium leguminosarum bv.phaseoli strains with which symptoms of N deficiency were notvisible (‘precocious’ strains). Cultivar Negro Argelwas then inoculated with two ‘traditional’ strains(C-05 and CIAT 727) and two ‘precocious’ strains(CNPAF 146 and CNPAF 512), and plants were harvested from 8to 30 DAE. There were no differences between the two groupsof strains in nodule dry weight or in the acetylene reductionrates between 8 and 16 DAE. However, nodules induced by the‘precocious’ strains showed earlier onset of glutaminesynthetase (GS) (EC 6.3.1.2 [EC] ) and glutamate synthase (GOGAT)(EC 1.4.1.14 [EC] ) activities, and ureide synthesis. The N concentrationin the nodules formed by ‘precocious’ strains variedfrom 4.2 to 4.5%, whereas with the ‘traditional’strains, it increased from 3.2% at 8 DAE to 65% at 18 DAE, atwhich time plants exhibited N-deficiency symptoms. By 21 DAE,GS and GOGAT activities in ‘traditional’ noduleswere increased, as well as the ureide-N-concentration in thexylem sap, nodule N content declined to 4.5% and the leavesbecame green. These results suggest that the N stress with ‘traditional’strains is not a limitation in early N² fixation activity butrather in the rates of expression of the processes of N assimilationand transport. Key words: Glutamate synthase, glutamine synthetase, nitrogen fixation, Phaseolus vulgaris, Rhizobium     

ERRATA

Effects of coupled solute and water flow in plant roots withspecial reference to Brouwer's experiment. Edwin L. Fiscus. p. 71 Abstract: Line 3 delete ‘interval’ insert‘internal’. p. 73 Materials and Methods: line 6: delete ‘diversion’ insert ‘division’ line 9 equation should read J_v=L_p P–RT(C⁰–C¹). 74 Last line of figure legend: 10^–1 should read 10^–11. 75 Line 11: delete ‘seems’ insert ‘seem’. le 1 column heading—10⁶ should read 10¹¹. 77 delete ‘...membrane in series of...’ insert ‘membranein series or...’ Delete final paragraph.     

Flagellate grazing on bacteria in a small dystrophic lake

Fluorescent beads were used to determine the grazing on bacteria by heterotrophic and mixotrophic flagellates in a highly humic (water colour 300–600 mg Pt l^–1) small lake. In summer phagotrophic flagellates constituted about three quarters of the numbers of phytoplankton (including heterotrophic or mixotrophic flagellates) in the uppermost epilimnion. Due to their small size their respective contribution to the biomass was about one quarter. The most important phagotrophic species were Ochromonas sp., and Chromulina spp. which ingested 75–203% of their body carbon per day from bacteria.In view of the abundance and biomass of phagotrophic and mixotrophic flagellates and their very high growth potential, they clearly play a significant role in the food chains of this lake. In addition to providing energy, bacteriovory also represents an important supply of inorganic and organic nutrients under nutrient limiting conditions.     

Pyrimidine Nucleotide Biosynthesis in Vinca rosea Cells: Changes in the Activity of the de novo and Salvage Pathways during Growth in a Suspension Culture

Marked changes in the activity of the ‘de novo’and ‘salvage’ pathways of pyrimidine biosynthesisduring growth of Vinca rosea cells in a batch suspension culturewere observed. The activity of these pathways was investigated by determiningthe contribution of ¹⁴C of [2-¹⁴Cluracil, 12-¹⁴Cluridine. and[6-¹⁴Clorotate to the cell constituents and by measuring theactivity of the several enzymes of these pathways. During the lag phase of the culture, ‘uracil-’ and‘uridine-salvage’ pathways made the predominantcontribution to nucleotide biosynthesis, but, following theinitiation of cell division, the ‘de novo’ pathwayfor nucleotide biosynthesis operated appreciably. These results suggest that nucleotide synthesis during cellgrowth in a suspension culture can be divided into two stages:a ‘turnover stage’, during the lag phase of cellgrowth, and a ‘true biosynthetic stage’, which isinitiated in the cell division phase.     

Remote detection of algae by copepods: responses to algal size, odors and motility

Selective feeding on large algae by copepods involves remotedetection of individual particles and subsequent active captureresponses In this study we use radiotracer experiments to quantifythe clearance rates of five coexisting freshwater copepods andto investigate the relative merits of the chemoreception andmechanoreception hypotheses of remote detection Tropocyclopsand three diaptomid copepods exhibited relatively high clearancerates when feeding on low concentrations of large algae, suggestingthat most previous studies with freshwater copepods have underestimatedmaximal clearance rates and the degree of size selectivity.All five species of copepods exihibited strong selection foran intermediate-sized flagellate (25 µm Cartena) or alarge-sized nonmotile alga (80 µ.m Pediastrum) over asmall-sized flagellate (6 µ.m Chlamydomonas). The weight-specificclearance rate for Tropocylops prasmus feeding on motile Cartena(271 ml mg^–1 h^–1) was about twice that of threediaptomid copepods and more than an order-of-magnitude higherthan the estimate for Epischura lacustris feeding on its preferredalga, Pediastrum Assuming that distance chemoreception (‘smell’)is important in remote detection, we predicted that the additionof high concentrations of ‘algal odors’ would obscureany chemical gradients emanating from individual algal cellsand would thereby hinder the remote detection and active captureof large algae Contrary to this hypothesis, the addition ofamino acids, sucrose, and algal extracts had no effect on theclearance rates and selectivity of Diaptomus birgei. These results,together with recent cinematographic studies (Vanderploeg etal.,1990), suggest that mechanoreception is the primary mechanismfor the remote detection of large particles by diaptomid copepods.A raptorial cyclopoid, Tropocyclops prasinus, exhibited strongpreferences for motile algae, whereas a suspension-feeding calanoid,D birgei, did not select between motile and nonmotile cells.Motility appears to be an important factor in algal detectionfor small cyclopoid copepods but not for suspension-feedingdiaptomids     

Impacts of nutrients and zooplankton on the microbial food web of an ultra-oligotrophic lake

In ultra-oligotrophic lakes and the sea, calanoid copepods arethe dominant mesozoo-plankton and cladocerans are generallysparse or absent. To determine the effects of predation andnutrient enrichment on the pelagic microbial food web of anultra-oligotrophic lake, we added copepods and cladocerans atlow biomasses (<60 µg l^–1 to in situ enclosuresin Lake Wakatipu, New Zealand, in the presence and absence ofadded nutrients (nitrogen and phosphorus). In response to nutrientfertilization, the concentrations of phototrophs >3 µmand heterotrophic bacteria increased by 50 and 15%, respectively,over 4 days, but those of cyanobacterial picoplankton decreasedby 68%. The presence of calanoid copepods (Boeckella dilatata)at ambient densities (1 and 4 l^–1) rapidly and severelysuppressed ciliate population growth over 4 days and also loweredthat of flagellates >3 µm, even when microbial growthwas enhanced by added nutrients. The presence of a small cladoceran,Ceriodaphnia dubia, at double the densities, but similar biomasses,to those of copepods, depressed the net growth rates of ciliatesand flagellates to a lesser degree. The net growth rate of heterotrophicbacteria after 4 days declined with flagellate abundance, consistentwith the possibility of regulation by flagellates. Althoughbacteria and algae increased in response to nutrient fertilization(bottom-up control), predation (top-down control) appeared toplay an important role in structuring the microbial food webof this ultra-oligotrophic lake in summer.     

The Sensitivity of Net Photosynthesis in Several Plant Species to Short-term Fumigation with Sulphur Dioxide

The effects of exposure of up to 2 h with sulphur dioxide ona range of plant species was observed by measuring changes inthe rate of net photosynthesis under closely controlled environmentalconditions. Ryegrass, Lolium perenne ‘S23’ was thespecies most sensitive to SO₂; significant inhibition was detectedat 200 nl l^–1. Fumigations at 300 nl l^–1 also inhibitedphotosynthesis in field bean (Vicia faba cv. ‘Three FoldWhite’ and ‘Blaze’) and in barley (Hordeumvulgare cv. ‘Sonja’). No effect was detected inwheat (Triticum aestivum cv. ‘Virtue’) at concentrationsup to 600 nl l^–1 SO₂, or in oil-seed rape (Brassica napuscv. ‘Rafal’) except at 800 nl l^–1 SO₂). Recoverycommenced immediately after the fumigation was terminated andwas complete within 2 h when inhibition had not exceeded 20%during the SO₂ treatment. Key words: Sulphur dioxide, short-term fumigation, photosynthesis     

Effect of Inorganic Phosphate on the Biosynthesis of Purine and Pyrimidine Nucleotides in Suspension-Cultured Cells of Catharanthus roseus

The levels of purine and pyrimidine nucleotides in suspensioncultures of Catharanthus roseus were determined 24 h after stationary-phasecells were transferred to fresh complete (‘+Pi’)or phosphate-deficient (‘–Pi’) Murashige-Skoogmedium. The levels of ATP, GTP, UTP and CTP were from approx.3 to 5-fold greater in the cells grown in ‘+Pi’medium than in the cells grown in ‘–Pi’ medium.The levels of almost all other nucleotides were slightly higherin the cells in ‘+Pi’ medium. The rates of de novoand salvage biosynthesis of purine and pyrimidine nucleotideswere estimated from the rates of incorporation of radioactivityfrom [¹⁴C]formate, [2–¹⁴C]glycine, NaH¹⁴CO₃, [6–¹⁴C]orotate,[8–¹⁴C]adenine, [8–¹⁴C]adenosine, [2–¹⁴C]uraciland [2–¹⁴C]uridine. The results indicated that the activityof both the de novo and the salvage pathway was higher in thecells in ‘+Pi’ medium than in the cells in ‘–Pi’medium. The rate of degradation estimated from the rate of releaseof ¹⁴CO₂ from labelled purines and pyrimidines indicated thatdegradation of uridine was significantly reduced in the cellsin ‘+Pi’ medium, but no significant difference wasfound in the degradation of adenine, adenosine and uracil. Thepossible role of Pi in the control of the biosynthesis of nucleotidesand in the degradation of uridine is discussed. Catharanthus roseus, Madagascar periwinkle, suspension culture, inorganic phosphate, nucleotides, purines, pyrimidines, biosynthesis, degradation     

Determinate Floral Buds of Plantain (Musa AAB) as a Site of Adventitious Shoot Formation

Floral buds of the ‘False Horn’ plantain clonesMusa (AAB) ‘Harton Verde’, ‘Harton Negra’,and ‘Currare’ terminate in a large single floralstructure. The apices of these floral buds are here designatedas determinate since they have lost the ability to produce additionalfloral initials or buds. Terminal peduncle segments can be culturedin a modified Murashige and Skoog (1962) medium supplementedwith N⁶-benzyl-aminopurine (5 mg I^–1). Under these conditions,this apparent inability to yield buds can be overcome as vegetativeshoot clusters form in the axils of the bracts. Rooted plantletsare obtainable by treating shoots with naphthaleneacetic acid(1 mg I^–1) and activated charcoal (0.025%). The adventitiousorigin of the shoots has been established. Musa cultivars, plantains, floral bud, adventitious buds, tissue culture     

Structure and dynamics of zooplankton communities maintained in closed systems, with special reference to the algal food supply

Seasonal changes in the structure of the zooplankton communitiesof two large (18 500 m³) experimental enclosures (‘LundTubes’) are reviewed. The diets of the two most prominentcrustaceans, Daphnia hyalina and Diaptomus gracilis, were investigatedby direct analyses of gut contents and in a series of shortterm experimental manipulations of communities isolated in 6m³ bags, placed in one of the enclosures. The relative abundanceof Daphnia hyalina generally coincided with warmer water temperaturesbetween late spring and late summer, but its numbers fluctuatedwithin that period, apparently in response to the availabilityof smaller algae (<10⁴ µm³ unit volume: chiefly cryptomonads,chlorellids and Ankyra). Daphnia numbers declined steadily throughthe summer in the enclosure overwhelmingly dominated by Microcystis,although it did feed on smaller colonies (< 10⁵ µm³in volume). Diaptomus gracilis and most rotifers were also prominentin the first half of the year, but peak abundances of Keratellaquadrata followed the presence of large populations of small(<10³ µm³) algae. Under conditions of limiting foodavailability, mortality was greatest amongst the smaller speciesand the juvenile instars of the large Crustacea. ¹Present address: Anglian Water Authority, North Street, Oundle,Peterborough, UK     

Relative Sensitivity and Tolerance of some Gladiolus Cultivars to Sulphur Dioxide

Five Gladiolus cultivars, namely ‘Aldebaran’, ‘BrightEye’, ‘Illusion’, ‘Manisha’ and‘Manmohan’, were exposed to 1 and 2 µg l^–1sulphur dioxide to test their relative-sensitivity toleranceto the pollutant Plants were fumigated experimentally for 2h daily Foliar injury symptoms were observed first in ‘Manisha’followed by ‘Aldebaran’ and ‘Illusion’at the higher dose Photosynthetic pigments and leaf extractpH were significantly decreased, particularly in ‘Manisha’and ‘Illusion’ Overall disturbances in the plantmetabolism due to SO₂ treatment led to retarded growth of plants,as evident from decreased shoot length and phytomass valuesThe order of sensitivity of the five Gladiolus cultivars toSO₂ was as follows, with the greatest first Manisha, Illusion,Aldebaran, Bright Eye, Manmohan Cultivars, Gladiolus, sensitivity, sulphur dioxide, tolerance     

Relative impacts of copepods, cladocerans and nutrients on the microbial food web of a mesotrophic lake

Calanoid copepods, rather than cladocerans, frequently dominatethe zooplankton of lakes in New Zealand. The potential consequencesof this domination for the microbial community of mesotrophicLake Mahinerangi, New Zealand, were determined by field experimentsin which Boeckella and Daphnia were added to in situ enclosuresin the presence and absence of added nutrients. Boeckella hamataat ambient densities (2 and 81^–1) rapidly and severelysuppressed ciliate population growth over 4 days, even whenmicrobial growth was enhanced by added nutrients, but effectsof copepods on other components of the microbial community (bacteria,photosynthetic picoplankton, heterotrophic nano-flagellates,algae) were slight. In contrast, Daphnia carinata at the samedensities (but 3-fold higher biomasses per litre) had a relativelyweak effect on ciliates, suppressing ciliate abundance onlyafter 4 days at 8 Daphnia 1^–1 (330 µg 1^–1);this daphniid density also depressed abundances of large bacterialrods, some photosynthetic picoplankton and the dominant alga,Cyclotella. These results highlight the relative importanceof specific trophic linkages in a microbial food web; they alsosuggest that the dominance of Boeckella in many southern hemispherelakes may account for relatively low ciliate abundances in theselakes.     

Behavioral responses of the brittle star Ophiura ophiura to amino acids, acetylcholine and related low-molecular-weight compounds

Feeding behavior of the brittle star Ophiura ophiura includesorienting posture, orienting movements, arm ‘walking’,changing the direction of ‘walking’ arm coilingand ingestion. All sequential behavior patterns were releasedor enhanced by single low-molecular-weight compounds. Stimuliwhich released ‘walking’ behavior at high concentrations(10^–4 M) in all the test animals are listed in decreasingorder of sensitivity: sarcosine, glycine, urea, L-valine, L-leucine,L-methionine, L-homocysteine, L-norvaline, L-norleucine, L-threonine,L-serine, S-methyl-L-cysteine, L-proline. Threshold values forsingle amino acids were as much as 100 times different in differentindividuals and ranged from 3 x 10^–9 to 3 x 10^–7M for the most effective stimulus, sarcosine, and from 10^–6to 10^–4 M for proline. Above 10^–5 M, only L-prolineregularly released a second behavior pattern, the arm coilingresponse, which temporarily inhibited the ‘walking’behavior. Behavioral thresholds for the ‘walking’behavior for L(+)-lactate and L-alanine were higher than thosefor the orienting movements. Thyoglycolic acid and ß-alaninereleased tube feet walking, which is not part of the feedingbehavior. Structure—activity comparisons were studied at estimated10^–5 M concentrations. Gycline, sarcosine, L-valine, L-norvaline,L-leucine, L-isoleucine, DL-norleucine and DL-homocysteine releasedarm ‘walking’ behavior in more than 75% of all thetests. With the exceptions of S-methyl-and S-ethyl-cysteine,and glycine methylester, derivatives of amino acids were noteffective behavioral stimuli in Ophiura ophiura. L-Isomers ofvaline and leucine regularly stimulated the ‘walking’behavior while their D-isomers were effective in some testsand ineffective in others. Acetylcholine iodide, acetyl-ß-methylcholine chloride and choline phosphate chloride regularly released‘walking’ behavior at concentrations above 10^–5M.     

Growth and mortality of flagellates and non-flagellate cells of Phaeocystis globosa (Prymnesiophyceae)

Two cell types of the same clone of Phaeocystis globosa, solitarynon-flagellate cells and flagellates, were grown in batch culturesunder identical conditions. The non-flagellate cells had a shorterlag phase (1.4 versus 2.8 days) and a higher growth rate (0.72versus 0.65 day^–1) than flagellate cells. The flagellateshad a longer stationary phase (15.6 versus 9.5 days) and a lowerdeath rate (0.07 versus 0.52 day^–1) than non-flagellatecells. All differences were statistically significant. Biomassyield did not differ between the two cell types. The short lagphase and high growth rate of nonflagellate cells correspondsto field observations of rapidly developing non-flagellate Phaeocystisblooms that are typically observed in nutrient-rich environmentssuch as temperate seas in spring. The flagellate cell type,with its longer stationary phase and lower death rate than non-flagellatecells, is better equipped for survival in oligotrophic environments.This explains why the flagellates of Phaeocystis are abundantafter the spring phytoplankton bloom in temperate seas and inother nutrient-poor environments such as the open ocean.     

Infection of Coscinodiscus spp. by the parasitoid nanoflagellate Pirsonia diadema: II. Selective infection behaviour for host species and individual host cells

The parasitoid nanoflagellate (PNF) Pirsonia diadema is hostspecific for the marine centric diatom Coscinodiscus spp. Experimentsshowed that flagellates significantly prefer C. wailesii overC.granii as host species (interspecific selectivity). This preferencewas independent of light conditions (dark, irradiance of 10and 70 µmol m^–2 s^–1) and temperature (10 and15C). Among unicellular host diatoms, the infection behaviourwas selective for individual cells: already infected C.graniicells were more attractive for further flagellate attachmentthan non-infected cells (intraspecific selectivity). Individualcells (     

Top-down effects of crustacean zooplankton on pelagic microorganisms in a mesotrophic lake

A series of single-factor in situ experiments was conductedin a mesotrophic lake in Brandenburg, North Germany, to studythe predatory impact of Eudiaptomus graciloides (adults, copepodites,nauplii), cyclopoid copepods (adult Diacyclops bicuspidatus,Thermocyclops oithonoides) and daphnids (adult Daphnia hyalina,Daphnia cucullata) on the microbial community (bacteria, autotrophicpicoplankton, flagellates, ciliates). All zooplankton speciestested reduced the ciliate community significantly and ingestionrates were always higher for ciliates in the 20–55 µmsize category as compared to smaller ciliates (10–20 µm).Adult E.graciloides, which exhibited the highest predatory impacton ciliates, differed from cyclopoids and daphnids by theirability to decimate ciliates to very low abundances. Ingestionrates of ciliates by the crustacean zooplankton followed thesequence E.graciloides > daphnids = cyclopoids = copepodites.While top-down control was evident for ciliates, top-down effectsdown to the autotrophic picoplankton and flagellates were mostlyrestricted to Daphnia-dominated treatments. Top-down effectswere never strong enough to produce negative bacterial growthrates. For all zooplankton tested, clearance rates for ciliatesexceeded those for phytoplankton. Besides the potential of thecrustacean zooplankton to influence the structure of ciliatecommunities, ciliates may contribute to the energy demands ofcopepods and daphnids, especially when phytoplankton resourcesare limited.     

Phytoplankton composition and cell carbon distribution in Prydz Bay, Antarctica: relation to organic particulate matter and its {delta}13C values

In January-February 1991, in Prydz Bay, phytoplankton bloomwas evident in the inner shelf area with the dominant diatomsbeing represented mainly by pennate species of the Nitzschia-Fragilariopsisgroup. Dinoflagellates and naked flagellates were most abundantin the centre of the bay; however, larger heterotrophic speciesprevailed at the southern stations. Cell carbon values (average317 µg l^–1; range 92-1048 µg l^–1) foundin the bloom in the south were chiefly due to pennate diatomsand larger heterotrophic dinoflagellates. Much lower carbonvalues (average 51 µg l^–1; range 7-147 µgl^–1) in the outer shelf region were mainly contributedby large centric diatoms (70-110 mu;m) and small dinoflagellates(5-25 µm). Wide ranges of algal cell sizes were observedin both southern and northern communities; the overlapping ofsizes of diatoms and flagellates, the latter containing heterotrophs,suggested complex trophic relationships within the planktonand an enhanced heterotrophic activity in the south. North-to-southvariations in surface     

Anther Culture of Lolium temulentum, Festuca pratensis and Lolium x Festuca Hybrids. I. Influence of Pretreatment, Culture Medium and Culture Incubation Conditions on Callus Production and Differentiation

This study was conducted with Lolium temulentum, Festuca pratensis,and the two hybrids L. multiflorum x F. pratensis ‘Elmet’and L. perenne x F. pratensis ‘Prior’. In a comparisonof various durations (7–42 d) of pretreatment at 4 or7 °C the highest yield of microspore-derived callus of L.temulentum was obtained after pretreatment of spikes at 7 °Cfor 28 d, conditions which also proved optimal for panicle pretreatmentwith F. pratensis. For ‘Elmet’, durations of 21–42d were optimal, and for ‘Prior’ the responses tendedto decline with increasing duration. In L. temulentum addition of charcoal (1–2 g l^–1)to medium containing 2, 4-D and KN wa