Immunohistochemical detection of opsins in turbellarians

Using the antiserum against the C-terminus of opsin of Dugesia japonica, immunohistochemistry revealed that opsins were actually localized in the photoreceptive organs of several turbellarians. The antiserum produced positive reactions in eyes of the Polycladida Notoplana koreana as well as those of the Tricladida Dugesia dorotocephala, Bdellocephala brunnea, Phagocata kawakatsui, Phagocata vivida and Polycelis auriclata, but not to those of Convoluta naikaiensis, Stenostomum sp. and Macrostomum tuba. This suggests that these opsin proteins of triclad and polyclad species have functional similarity in the C-terminus. This revised version was published online in July 2006 with corrections to the Cover Date.     

Parvalbumin-immunoreactive neurons in the brain of Helix pomatia

Parvalbumin-immunoreactive material was detected in the central nervous system of the snail, Helix pomatia. Each ganglion investigated contained parvalbumin-immunoreactive neurons. The molecular weight of Helix parvalbumin-immunoreactive material as determined by Western blots is about 40 kilodaltons. ⁴⁵Ca²⁺ overlays showed that this protein binds Ca²⁺. In contrast to vertebrates, in Helix neurons parvalbuminlike material was not colocalized with the neurotransmitter gamma-aminobutyric acid (GABA).     

Studies on the speciation of the European freshwater planarians Polycelis nigra and Polycelis tenuis based on the analysis of enzyme variation by means of iso-electric focusing

Polycelis nigra Ehrenberg and Polycelis tenuis Ijima differ morphologically and karyologically. No difference, however, was found in the isozyme pattern of malate dehydrogenase and tetrazolium oxidase, indicating a close relationship. Most sibling species differ at half of the loci.It could be deduced that the reproductive behaviour of a single population of Polycelis nigra in a Dutch pond was not panmictic. Two genetically different strains retained their identities during two years of observation. If pseudogamy occurs in this diploid planarian, the presence of heterozygous specimens indicates the absence of a true meiosis.The iso-electrofocusing technique by which these population-genetical studies were carried out, also lends itself to a comparison of overall protein banding patterns. The membrane proteins especially are conservative. The sodium dodecyl sulphate extracted proteins of Polycelis nigra-tenuis, Planaria torva and Phagocata vitta were very similar, while their water soluble proteins were not. This technique may be of great help in taxonomic studies of the higher taxa.     

Characterization of small GTP-Binding proteins in plant cell

To identify and characterize small GTP-binding proteins in plant cells, GTP-binding studies were performed with electroblotted plant proteins following SDS-polyacrylamide gel electrophoresis using [α-³²P]GTP. Three species of small GTP-binding protein (21, 23, and 27 kD) which have a specific GTP-binding property were identified in the membrane and cytosolic fractions of both monocotyledons (Zea mays) and dicotyledons (Glycine max). Moreover, these three species of small GTP-binding protein were gradually decreased when membranes were treated with hydroxylamine. This result indicates that these small GTP-binding proteins in plant cells are fatty acylated to the membrane lipids. The 27 kDa component was partially purified from hypocotyl membranes of Glycinemax, following S-300 gel filtration, phenylsepharose CL-4B, hydroxyapatite, and Q-sepharose column chromatography. This 27 kD protein was found to have both GTP-binding and GTPase activities.     

Abundanzdynamik und reproduktionsphasen limnophiler tricladen (Turbellaria) Aus stehenden kleingewässern

Dynamics of abundance and reproductive cycles of limnophileous triclads (Turbellaria) from little ponds.

1. Studying 18 little ponds in Lower Southern-Saxonia (West-Germany), in 14 ponds seven triclad-species were found.
2. Phagocata vitta occurs from October to July in 1–2 generations. In the area the species reproduces by fissipary. Maximum abundances and rates of reproduction are reached in December and January. In the ponds, where P. vitta and Dugesia polychroa live together, there is no competition between the two species.
3. After dry periods Dendrocoelum hercynicum emigrates from interstitial habitats as facultative inhabitant of surface-waters.
4. In low abundances Dendrocoelum lacteum lives in one pond only. The breeding period (production of cocoons) lasts from January to March. Low densities of this species are probable caused by interspecific food-competition with Polycelis nigra.
5. Likewise, Dugesia tigrina inhabits only one pond. The species is competitive to P. nigra at temperatures of about 20°C. High abundances in the months July to October fall together with high fissipareous-rates.
6. Dugesia polychroa occurs in low densities over the year or dependent on dry-periods. Cocoons are produced between March and May, in low numbers till autumn.
7. In the stagnant pond Bursfelde Polycelis nigra is the absolute dominant triclad-species with densities of up to 800 individuals/0,I m³. The maximum-abundances are caused by two intense reproductive periods in spring and autumn, together with optimum temperatures and food conditions.

     

Benthic invertebrates of Rutland Water

The colonisation of Rutland Water by benthic invertebrates has been monitored from 1976 when the reservoir was first filled. A modified Ekman grab was used to sample populations quantitatively while false substrates and dredge samples were used to obtain supplementary data. Hand net and hand-sorting were used to study the littoral zone.The dominant taxa in the reservoir are Asellus aquaticus, A. meridianus, Gammarus pulex, Crangonyx pseudogracilis, Helobdella stagnalis, Erpobdella octoculata, Polycelis tenuis, larval Chironomidae and Oligochaeta. Other species which appeared abundantly, such as Limnaea pereger and Planorbis albus, have now disappeared and it is suggested that the generally low levels of gastropods might account for the poor performance of species such as Dugesia lugubris and Glossiphonia complanata.Other aspects of abundance are discussed, as are the effects of maturation of the system.     

Regeneration of freshwater planarians <Emphasis Type="Italic">Dugesia tigrina</Emphasis> and <Emphasis Type="Italic">Polycelis tenuis</Emphasis> under the influence of methyl mercury compounds of natural origin

We studied the effects of methyl mercury compounds of natural origin on regeneration of the planarians Dugesia tigrina and Polycelis tenuis. Accumulation of methyl mercury in the planarian body leads to a delayed formation of photoreceptor organs in planarians of both species. After a significant traumatic load, the regeneration is suppressed and the death of some control and most experimental animals was observed. The intensity of joining additional cuts depends on the localization of body fragment with a cut and localization of a cut itself.Translated from Ontogenez, Vol. 36, No. 1, 2005, pp. 35–40.Original Russian Text Copyright © 2005 by Medvedev, Komov.     

An immunocytochemical method for histamine: application to the planarians

Histamine-immunoreactivity was investigated in the planarians Dugesia tigrina and Polycelis nigra. Specific antisera against a histamine-protein conjugate were used, and 1-ethyl—3 (3-dimethyl-aminopropyl) carbodiimide was used both as coupling agent to prepare the antigen and as a tissue fixative. In D. tigrina, histamine-immunoreactivity was restricted to photoreceptor cells in the cerebral eye. In P. nigra, nerve fibers were found in the ventral nerve cord and nerves running laterally from these. The epidermal eyes did not display histamine-immunoreactivity. The results suggest that histamine may be a transmitter in some of the most primitive animals. They also suggest that the distribution of histamine may differ in planarians.     

Clonal variation in apical growth and content in vegetative storage proteins in Populus

Summary Apical shoot growth and storage protein content in various poplar species and clones were followed in trees growing in the field and in micropropagated plants cultivated in the growth chamber under a controlled environment. In autumn a 32 kD and a 36 kD vegetative storage protein accumulate in wood, bark and roots of poplar and comprise together about 25% of the soluble proteins. In spring, at the time of dormancy break, the storage proteins are degraded and 3 weeks after budburst these proteins are no longer immunologically detectable. As in autumn, short day exposure of black cottonwood plants (Populus trichocarpa Torr. and Gray) induces cessation of apical growth and accumulation of the 32 kD and 36 kD vegetative storage proteins in all clones studied. In order to simulate spring conditions, short day induced plants were transferred back to long days. Like the situation in spring, budburst and storage protein degradation occurred considerably earlier in clone 9/60 than in clone Muhle Larsen. The latter clone accumulates both in winter and after short day exposure more storage proteins than the former. Furthermore two P. trichocarpa clones differ qualitatively in storage protein content: they possess an additional 34 kD polypeptide which cross-reacts with the anti-32 kD antibody. In conclusion, apical shoot growth and the capacity to synthesize storage proteins can be easily followed in micropropagated poplar cultivated in the growth chamber under inducing photoperiods. This offers the major advantage of independence from the annual growth cycle. Within one species considerable clonal variance in storage protein content and in the induction times needed for dormancy and dormancy break were observed. The suitability of storage protein content and apical growth as early selection traits in breeding programs focusing on nitrogen efficient poplar and clones adapted to specific latitudes will be discussed.     

Regulation of Phosphorylation of Wheat Mitochondrial Proteins by Divalent Cations

Mitochondria isolated from 4-day-old dark-grown wheat seedlings were purified by self-generating Percoll gradient. Phosphorylation reaction was carried out in vitro with the addition of [ c-³²P]ATP and polypeptides resolved by 50S-PAGE were subjected to autoradiography. Amongst endogenous polypeptides phosphorylated, four polypeptides of 120, 66, 43 and 21 kD were prominent. Addition of Mg²⁺ (5 mM) caused dephosphorylation of 120 and 66 kO polypeptides but, simultaneously, induced/enhanced the phosphorylation of some polypeptides, with the effect being more pronounced on a 67 kD species. The phosphorylation of 120 kD species and a few other polypeptides was also down-regulated and that of a 18 kD polypeptide was up-regulated by Ca²⁺. The present study provides evidence that phosphorylation status of mitochondrial proteins is regulated by Mg²⁺ and/or Ca²⁺-dependent phosphatase(s) and protein kinase(s).     

Continuing dispersal of freshwater triclads (Platyhelminthes; Turbellaria) in Britain with particular reference to lakes

1. A survey of lakes in Anglesey and Snowdonia, Wales, in the summer of 1997 recorded changes in the triclad fauna which had occurred since earlier studies in the 1950s and in 1973. Two native species, Polycelis tenuis and Dugesia polychroa, and two immigrants, Planaria torva and especially Dugesia tigrina, have increased their range. The triclad fauna has remained unchanged with time in eleven out of sixteen lakes in Snowdonia, in contrast to only one out of fourteen lakes on Anglesey. This supports the hypothesis produced in the 1950s, that the triclad fauna of the ion-poor, unproductive lakes in Snowdonia would change little with time, being maintained by ecological interactions, whereas the ion-rich, productive lakes on Anglesey, from which native Dugesia spp. and Dendrocoelum lacteum are historically absent, would increase in triclad diversity. 2. An examination of national records for the distribution of D. tigrina and Pl. torva indicated their continuing dispersal in both still and running waters in mainland Britain, with the former species having the greater frequency of occurrence. 3. The reasons for this may be that D. tigrina is (a) more easily dispersed, (b) a more opportunistic, catholic feeder, (c) a more vigorous competitor, (d) able to reproduce asexually and (e) able to prey to some extent on other triclad species, as shown in the laboratory but not yet confirmed in the field. The absence of D. tigrina and Pl. torva from unproductive lakes may be explained in terms of low temperature and feeding mode, and a low standing crop of gastropods, respectively. 4. Seven surveys (1961–1997) of the triclad fauna (six species) of Colemere, England, revealed that D. tigrina was confined to one small area of the littoral zone until the mid-1980s, after which it spread fairly rapidly to occupy the entire shore by 1997. Its expansion has been to the detriment of all the native species, particularly P. tenuis, except Dd. lacteum. The possible extinction of Pl. torva may be linked to a numerical reduction in snails, particularly Potamopyrgus jenkinsi. 5. Dugesia tigrina has an adverse effect on the native triclad fauna in lakes, particularly when only a few species are present. This has been explained in terms of the availability of a wider variety of food. The presence of a larger number of triclad species denies the immigrant food items, particularly crustaceans and snails, the food refuges of Dd. lacteum and Dugesia spp., respectively. However, the long-term study of Colemere has shown that more diverse triclad faunas may also be vulnerable to this immigrant. The effect of Pl. torva on native triclads remains uncertain.     

The identification of a heat-shock protein complex in chloroplasts of barley leaves

In vivo radiolabeling of chloroplast proteins in barley (Hordeum vulgare L. cv Corvette) leaves and their separation by one-dimensional electrophoresis revealed at least seven heat-shock proteins between 24 and 94 kD, of which most have not been previously identified in this C₃ species. Fractionation into stromal and thylakoid membrane components showed that all chloroplast heat-shock proteins were synthesized on cytoplasmic ribosomes, translocated into the chloroplast, and located in the stroma. Examination of stromal preparations by native (nondissociating) polyacrylamide gel electrophoresis revealed the presence of a high-molecular mass heat-shock protein complex in barley. This complex was estimated to be 250 to 265 kD in size. Dissociation by denaturing polyacrylamide gel electrophoresis revealed a single protein component, a 32-kD heat-shock protein. The synthesis of this protein and the formation of the heat-shock protein complex were dependent on functional cytoplasmic ribosomes. Immunological studies showed that the heat-shock protein complex did not contain any proteins homologous to the α-subunit of ribulose bisphosphate carboxylase oxygenase subunit-binding protein. Other features about the complex included the absence of nucleic acid (RNA or DNA) and its nondissociation in the presence of Mg²⁺/ATP. These results suggest that the heat-shock protein complex in barley chloroplasts is a homogeneous octamer of 32-kD subunits.     

Resistance of two planarian species to UV-irradiation

The aim of this work was to determine the effects of 20, 25 and 30 minute UV-irradiation periods lambda = 253.5 nm to two planarian species Dugesia tigrina (Gir.) and Polycelis felina (Daly.). In vivo, UV light effects have been reported to affect intracellular receptors and disrupt simple behaviour. The effects of UV-rays on mortality and behavior as well as morphological, cytological and histological changes in the two planarian species were assessed, and the course and the dynamics of regenerative processes were compared between them. Experimental populations of Dugesia tigrina and Polycelis felina species were maintained in laboratory conditions at room temperature. Mortality, behavioral and morphological changes were monitored daily by means of a light stereomicroscope. For cytological and histopathological analysis, planarians were fixed in Bouine fixative on the first, second, third, fifth and seventh day after exposure to UV-irradiation, respectively. They were embedded in paraffin, cut on a microtome, stained with toluidin blue and embedded in Canada-balsam. UV-rays caused mortality, behavioral, morphological, cytological and histological changes in each planarian species. In regeneration of damaged body parts reticular cells and neoblasts played the main role. Neoblasts as totipotent cells extremely increased in number in the area of damaged tissue, immediately after UV-exposure. Dugesia tigrina was more sensitive to UV-rays than Polycelis felina due to possession of less pigmented cells. The course of regeneration in both species was similar. Most individuals of both species regenerated in 5 to 12 days after UV-irradiation.     

DNA-Binding Proteins and Structural Characteristics of Chloroplast Nucleoids

A comparative analysis of proteins from chloroplast nucleoids was performed in two higher-plant species (Pisum sativumL. andArabidopsis thalianaL.) and a green alga Chlamydomonas reinhardtiiDang. In the nucleoids of the higher plants and the alga, 26–27 proteins were detected with their mol wts ranging from 10 to more than 94 kD. In all the species tested, the distribution of nucleoid proteins by their mol wts was similar, especially between the predominant proteins with mol wts of 10 to 40 kD. Six DNA-binding proteins (12–18 kD) were detected in nucleoids fromCh. reinhardtiichloroplasts after in vivocovalent cross-linking between chloroplast DNA and proteins. Under an electron miscroscope, some regular structures resembling nucleosome-like particles of bacterial cells were revealed.     

Observations on the comparative ecology of lake-dwelling triclads in southern Sweden,Finland and northern Britain

Summary There are more lake-dwelling species of triclad in northerm Britain (10) than in southern Sweden (7) and the distribution of some differ in the two countries over approximately comparable types of lake. Dendrocoelum lacteum (Müller) occurs in a wide range of lake types in s. Sweden but is restricted to more productive lakes in n. Britain. Polycelis nigra (Müller) (absent from Sweden) has a similar, wide occurrence in Britain. Planaria torva (Müller) is more common in s. Sweden in lakes of all types but particularly in the less productive lakes. The Polycelis species P. tenuis (Ijima) and P. hepta E. H. and Y. Melander, also the Dugesia lugubris species complex show much the same distribution in both areas. Polycelis felina (Dalyell) does not occur in Sweden. The small nuber of Finnish lakes sampled confirm the data for s. Sweden. There is significant correlation between the total abundance of triclads and both the hardness and dissolved matter contents of the Swedish lake waters. Quantitatively the s. Swedish triclad populations compare most closely with those of Mid-Scotland in the n. Britain area.

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Zusammenfassung Es gibt im Norden Britanniens eine grössere Anzahl von Tricladen (10) die Süsswasserseen bewohnen, als in Süd-Schweden (7) und einige Sorten verbreiten sich in ungefähr gleichen Arten von Seen in verschiedenem Maasse in den beiden Ländern. Dendrocoelum lacteum (Müller) erscheint in einer grossen Reihe verschiedenartiger Seen in Süd-Schweden, beschränkt sich aber im Norden Britanniens auf die Seen mit grösserem Reichtum an Lebewesen. Polycelis nigra (Müller) — in Schweden nicht vorhanden-, hat eine ähnlich weite Verbreitung in Britannien. In Süd-Schweden ist in Seen aller Art ein reicherer Bestand an PLanaria torva (Müller), besonders in den Seen mit geringerem Bestand an Lebewesen. Die Polycelis Art P. tenuis (Ijima) und P. hepta E. H. und Y. Melander und auch der Dugesia lugubris Arten-Komplex zeigen dieselbe Verbreitung in beiden Gegenden. Polycelis felina (Dalyell) kommt in Schweden nicht vor. Die kleine Zahl finnischer Seen, denen Proben entnommen wurden, bestätigen die Befunde von Süd-Schweden. Wesentlicher Zusammenhang besteht zwischen der gesamten Reichhaltigheit von Tricladen und dem Gehalt an Kalk und aufgelöster Substanz bei den schwedischen Süsswasserseen. Mengenmässig ist die Verbreitung von Tricladen in Süd-Schweden der im Norden Britanniens am echesten zu verleichen, soweit es sich um Mittel-Schottland handelt.

     

Chromosome bands in freshwater triclads

Four species of Triclads belonging to three families, Dugesia polychroa and Dugesia mediterranea (Dugesiidae), Planaria torva (Planariidae) and Dendrocoelum lacteum (Dendrocoelidae) were studied for chromosome banding: C-banding for all the species, ASG banding for Dugesia polychroa and G-banding for Dugesia polychroa and Dugesia mediterranea. C-banding results for Dugesia lugubris-polychroa group suggest a high level of heterochromatin evolution in the group. Small bands strictly limited to the centromeric region were seen in Dugusia lugubris and this pattern is similar to the patterns of some species belonging to other families, notably Planaria torva and Dendrocoelum lacteum. This could be considered a primitive character. In contrast numerous heterochromatin bands which are useful to characterize the different karyotypes were seen in Dugesia polychroa and Dugesia mediterranea. Our data suggest that heterochromatin is important in freshwater triclad speciation.     

Ginogenesi in tricladi di acqua dolce

Zusammenfassung Bei der Rasse G von Dugesia benazzii von Sardinien hat das Spermatozoon nur die Funktion der Entwicklungserregung, da die Amphimixis nicht erfolgt und der männliche Vorkern wie ein Eichtungskörper eliminiert wird.Das Ei entwickelt sich allein mit der weiblichen Chromosomengarnitur (Gynogenese), und das erklärt die vom Autor an dieser Planarie festgestellte mütterliche Vererbung.Auch bei den italienischen Rassen von Polycelis rigra erfolgt die Entwicklung durch Gynogenese nach den genetischen Untersuchungen des Autors und den cytologischen von N. G. Lepori.     

The consequence of a food refuge collapse on a guild of lake-dwelling triclads and leeches

The diets ofPolycelis nigra, P. tenuis, Dugesia polychroa, Dendrocoelum lacteum, Glossiphonia complanata, Helobdella stagnalis andErpobdella octoculata in an English lake were examined, using a serological technique, and compared between 1981–82 and 1989–90. Leech, triclad and prey abundances were also recorded. Between the two studies, snail numbers crashed whereas the abundances ofAsellus andGammarus increased.Dugesia andGlossiphonia numbers decreased substantially, whilstPolycelis tenuis andHelobdella abundances increased. In the second study, the snail component in the diet was greatly reduced resulting in a broader food niche, particularly forDugesia andGlossiphonia, and greater food overlap between the predators with the exception ofDendrocoelum andErpobdella which do not eat molluscs. It is postulated that the reduced size of the snail refuge, and consequent increase in severity of interspecific competition with other predators, particularlyPolycelis andHelobdella, led to the observed decrease in abundances ofDugesia andGlossiphonia. The decline in the last two genera, perhaps coupled with increased crustacean abundance, could have contributed to the numerical increase of the competitively superiorPolycelis tenuis andHelobdella.     

Changes of alkaline phosphatase activity in response to different stressors in planarian Dugesia japonica

The aims of this work are to provide some properties of alkaline phosphatase (ALP) in the planarian Dugesia japonica and detect its activity in response to different stressors, as well as to introduce renatured SDS-PAGE to study enzyme activity in planarians. Our results indicate that ALPs in planarians are mainly membrane-bound form, identified as three mainly enzyme-bands (approximately MW 260 kD, 180 kD, 160 kD, respectively). Under our experimental conditions, ALPs activity had no apparent changes in response to low concentration of Hg²⁺ (25 μg L^?1) and Pb²⁺ (125 μg L^?1, 250 μg L^?1) exposure, but were severely inhibited in response to high concentration of Hg²⁺ (50 μg L^?1, 150 μg L^?1, 300 μg L^?1) and Pb²⁺ (500 μg L^?1, 1000 μg L^?1) exposure. Mild heat shock (25°C for 2 days) elevated ALP activity, but severely heat shock (25°C for 2 days, followed by 30°C for 2 days and 32°C for 2 days) inactivated ALP activity. Interestingly, ALP and other cytosolic phosphatases (MW from ～45 kD to ～60 kD) activity increased noticeably during the early stage of planarians regeneration, which may be involved in cell proliferation and differentiation. Contrary to regeneration, prolonged starvation suppressed ALP activity. The above findings provide valuable information about the role of ALP in planarian regeneration and for its use as an indicator in ecotoxicology.     

Changes in Phosphorylation Status of Wheat Plastid Polypeptides as Influenced by Light, Calcium and Cyclic AMP

The polypeptides of etioplast and chloroplast fractions, purified on Percoll discontinuous gradient, were phosphorylated in vitro using (γ-³²P)ATP, resolved by SDS-PAGE and autoradiographed. In general, about 15-18 phosphopolypeptides in the range of 14-150 kD were distinctly visible in autoradiograms of both organelle fractions with varying degree of radiolabel incorporation. Although short-term irradiation with red or far-red light did not have any significant effect on phosphorylation status of etioplast polypeptides, in vivo irradiation with 1 h white light, followed by in vitro phosphorylation, decreased phosphorylation of a 116 kD polypeptide and increased the phosphorylation of polypeptides of 38 kD and a doublet around 20 kD. Strikingly, the phosphorylation status of 116 kD etioplast polypeptide was adversely affected by Ca²⁺ as well, and this phosphopolypeptlde was not distinctly visible in the autoradiogram of the chloroplast fraction proteins. However, in vitro phosphorylation of 98, 57 and 50 kD polypeptides of both etioplast and chloroplast fractions was found to be Ca²⁺ dependent. Unlike Ca²⁺, 3′,5′-cyclic AMP down-regulated the phosphorylation of several polypeptides of both etioplasts and chloroplasts, including 98 and 50 kD, and up-regulated the phosphorylation of 32 and 57 kD polypeptides. The significance of these observations on changes in phosphoprotein profile of etioplasts and chloroplasts, as influenced by light, Ca²⁺ and cyclic nucleotides, has been discussed.