Beobachtungen zur Lebensweise von Pycnogonum litorale (Ström) (Pantopoda)

Zusammenfassung Am Leitdamm des Jadebusens lebt Pycnogonum litorale im Lückensystem des Miesmuschelbesatzes. Dieser bietet mit hartem Untergrund, hoher Feuchtigkeit bei Niedrigwasser, genügend Actinien als Nahrung und guter Durchströmung bei gleichzeitigem Schutz vor Vertragung—offensichtlich günstige Lebensbedingungen für Pycnogonum litorale.Der Eiablage im Februar geht eine Reiterstellung des Männchens auf dem Weibchen von durchschnittlich 24 Tagen voraus. Unter künstlichen Kurztagbedingungen kann diese Reiterstellung auch außerhalb der Fortpflanzungsperiode eingenommen werden. Die Eier werden durch Rumpfbewegungen beider Partner zu den Ovigeren des Männchens bewegt. Bei 12°C schlüpfen die Larven etwa 41 bis 46 Tage nach der Eiablage aus, bei 19°C, im Sommer, schlüpften keine Larven.Im Jadebusen leben die Larven etwa 1/2 Jahn endoparasitisch in Hydrozoen. Die an die Metamorphose anschließende juvenile Phase, in der die Tiere frei leben, dauert ein knappes Jahr, die Reifehäutung erfolgt normalerweise im Sommer des zweiten Jahres, die Fortpflanzungsperiode etwa 6 Monate später, im Winter.

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Observations on the life biology of Pycnogonum litorale (Ström) (Pantopoda)

Summary Pycnogonum litorale lives in an interstitial system, of the mussel zone on the embankment of the Jadebusen. Hard substrate, high humidity at low tide, sufficient Metridium senile as food, and active currents together with protection from drifting, constitute favourable conditions for this pycnogonid.Prior to laying egg in February, the male remains in a riding position upon the female for approximately 24 days. Under artificial short-day conditions the riding position may also be assumed outside of the reproductive period. The eggs are transported to the ovigers of the male by trunk movements of both partners. At 12°C the larvae hatch about 41–46 days after egg-laying. No larvae hatched from eggs laid during summer at 19°C.The larvae live endoparasitically in Hydrozoa for about 1/2 year. Following metamorphosis, the freeliving juvenile phase lasts barely a year. The maturation moult normally takes place in the summer of the second year, the reproductive period beginning about 6 months later, in winter.

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Mit Unterstützung der Deutschen Forschungsgemeinschaft.     

Larval development and morphogenesis of the sea spider Pycnogonum litorale (Ström, 1762) and the tagmosis of the body of Pantopoda

Aspects of pantopod ontogeny have been known for a long time, but specific information is available for only a few species. Our account of the postembryonic development of Pycnogonum litorale is based on laboratory-reared individuals and SEM studies. We documented particularly all early developmental stages, with emphasis on morphogenetic changes of head structures and appendages. In P. litorale the protonymphal limbs, the chelicerae and two more uniramous legs, degenerate already during the larval phase; only the third one, the ovigers, reappears in male juveniles. Other Pantopoda vary in this aspect from retention of all three protonymphal appendages to their complete reduction, as in P. litorale. Accordingly, the two post-cheliceral larval appendages are separate legs in front of the walking legs in the adults, the ‘parapalps’ and the ‘ovigers’, but they do not occur in all pantopods. The scarcity of studies of the ontogeny of Pantopoda prevents us from a more conclusive picture, but our data are promising to state that additional such studies will increase the usability of ontogenetic data for a phylogenetic analysis of Pantopoda, the crown group of the Pycnogonida. We also discuss the phylogenetic implications of our data in the light of new information from Hox genes and developmental-biological data on body segmentation and tagmosis of the Chelicerata. These suggest the homology of chelicerae and antenn(ul)ae of other euarthropods. Accepting this, we conclude that the adult pycnogonid/pantopod head, the cephalosoma, corresponds to the euarthropod head and that the protonymph with three appendage-bearing segments may represent an even shorter, possibly phylogenetically older larval type than the euarthropod ‘head larva’ bearing four pairs of appendages. In further consequence, the fourth walking legs of Pycnogonida/Pantopoda should correspond to the first opisthosomal appendages, the chilaria, of euchelicerates. This implies that within Pycnogonida the post-prosomal region became compacted during evolution to a single leg-bearing segment plus a tubular end piece. Accordingly, neither the anterior nor the posterior functional boundaries of the walking-leg region correspond to the original tagma borders.     

Cleavage and gastrulation in Pycnogonum litorale (Arthropoda, Pycnogonida): morphological support for the Ecdysozoa?

The early cleavage and gastrulation of the pycnogonid Pycnogonum litorale is investigated in detail by fluorescence microscopy, confocal laser scanning microscopy, and histology. The cleavage is holoblastic with equally sized blastomeres and an irregular radial pattern. There is no stereotypic cell lineage, and timing and spindle directions of individual mitoses vary to a high degree. Gastrulation begins at the 63-cell stage with the retardation and enlargement of a cell which adopts the form of a bottle and fills the interior of the egg, followed by immigration and epiboly of smaller cells surrounding the large bottle-shaped cell. The gastrulation site marks the dorsal side of the embryo and the stomodaeum forms adjacent to the area of gastrulation. The pattern of the early development of Pycnogonum is compared with that of other Pycnogonida resulting in a putative ground pattern of pycnogonid development. Furthermore, our results are discussed in the wider framework of putative arthropod and cycloneuralian relationships. This comparison implies morphological support for the Ecdysozoa.     

Isolation and identification of α-proteobacteria from Culex pipiens (Diptera Culicidae) larvae

A survey of drainage ditches in suburban areas of La Plata, Buenos Aires province, Argentina for pathogens of Culex pipiens larvae was conducted from 2003 to 2006. C. pipiens larvae of opaque, white color were found in several of those field collections. When the white larvae were dissected and observed by phase-contrast microscopy in wet-mount preparations, the presence of bacteria, located in the hemocoel, was recorded. Laboratory experiments were performed to elucidate the pathway for transmission of this pathogen. Although approaches involving traditional culturing had failed to reveal the identity of the new microorganism present, molecular techniques to identify the pathogen in the studies reported here were successful. The partial sequence of the 16S-rRNA gene constitutes a powerful tool for the detection of new isolates from the hemocoele of C. pipiens larvae. These bacteria were characterized as belonging to the genus Novispirillum. In spite of the genus's wide distribution in different aquatic environments, information related to the parasitic relationship of Novispirillum spp. to aquatic insects is scarce, and this association has not been described in other mosquito species. This report constitutes the first documentation of Novispirillum spp. as a pathogen for mosquito larvae.     

Isolation and identification of entomopathogenic nematodes and their symbiotic bacteria from Hérault and Gard (Southern France)

Isolation and identification of native nematode-bacterial associations in the field are necessary for successful control of endemic pests in a particular location. No study has yet been undertaken to recover and identify EPN in metropolitan France. In the present paper, we provide results of a survey of EPN and their symbiotic bacteria conducted in Hérault and Gard regions in Southern France. Molecular characterization of isolated nematodes depicted three different Steinernema species and one Heterorhabditis species, H. bacteriophora. Steinernema species recovered were identified as: S. feltiae and S. affine and an undescribed species. Xenorhabdus symbionts were identified as X. bovienii for both S. feltiae and S. affine. Phylogenetic analysis placed the new undescribed Steinernema sp. as closely related to S. arenarium but divergent enough to postulate that it belongs to a new species within the “glaseri-group”. The Xenorhabdus symbiont from this Steinernema sp. was identified as X. kozodoii. All Heterorhabditis isolates recovered were diagnosed as H. bacteriophora and their bacterial symbionts were identified as Photorhabdus luminescens. Molecular characterization of these nematodes enabled the distinction of two different H. bacteriophora strains. Bacterial symbiontic strains of these two H. bacteriophora strains were identified as P. luminescens ssp. kayaii and P. luminescens ssp. laumondii.     

Isolation,identification and subtyping of Campylobacter: Where to from here?

Campylobacter species are widely regarded as the most frequent bacterial cause of gastroenteritis in humans worldwide. Their main transmission routes are via contaminated food and water. For interventions to be effective, methods for the detection, identification and epidemiological subtyping must be sensitive, accurate and rapid. As yet, methods are not perfect, although several significant advances have been made in these areas in recent years. This paper provides a brief review and commentary on the current state of the art in the hope that it will help provide context for others in selecting, improving or developing these vital tools for research and diagnoses.     

Characterization of the murine Lbx2 promoter, identification of the human homologue, and evaluation as a candidate for Alström syndrome

The murine Lbx2 gene is a member of the ladybird family of homeobox genes, which is expressed in the developing urogenital system, eye, and brain. Using transgenic mice, we demonstrate that 9 kb of the 5' flanking region of mouse Lbx2 is able to direct expression of a reporter gene in a tissue-specific manner recapitulating the endogenous expression pattern. This regulatory region provides a novel reagent allowing for transgenic expression in the developing urogenital ridge. In addition, we describe the identification of the human homologue, LBX2. Comparison of the human LBX2 and mouse Lbx2 sequences upstream of the coding regions reveals sequence conservation suggesting conserved regulatory regions. Both the human LBX2 and the mouse Lbx2 genes have similar genomic structures and are composed of two exons separated by an intron. We mapped the mouse Lbx2 gene to 35 cM on chromosome 6 and the human LBX2 gene to a homologous region of chromosome 2p13. This is a candidate region for several inherited disorders, including Alstr?m syndrome, a disorder that includes ocular, urogenital, and renal abnormalities. Given the expression pattern of Lbx2, the chromosomal location in humans, and the potential function of mammalian ladybird genes, we have begun to analyze patients with ocular disorders and those with Alstr?m syndrome for mutations in LBX2. Although polymorphisms were identified, our results indicate that mutations in the coding region of LBX2 do not account for Alstr?m syndrome in the six kindreds analyzed.     

Isolation and re-association of the subunits from the pro-(carboxypeptidase A)–pro-(proteinase E) binary complex from pig pancreas

The component subunits of the pro-(carboxypeptidase A)–pro-(proteinase E) binary complex from pig pancreas were separated with a high recovery (80–95%) of their original potential activity. The isolated subunits and the reconstituted complex have properties similar to those of the corresponding natural species. The tryptic activation course of the pro-(carboxypeptidase A) subunit is substantially modified when bound to pro-(proteinase E), whereas the activation of pro-(proteinase E) is not dependent on this association.     

A new species of Austrodecus Hodgson, 1907 (Arthropoda,Pycnogonida, Austrodecidae) from?the?Southwest Indian Ridge

A new species of pycnogonid collected by the Chinese research vessel R/V Dayangyihao during cruises to the Southwest Indian Ridge in 2008 and 2009 is recorded. The new species, Austrodecus bamberi, is placed into the tristanense-section by the characters of 4-articled ovigers and present auxiliary claws and is distinguished from other species in this section by the number and length of tubercles on the first coxae.     

Isolation and functional characterisation of the genes encoding Δ(8)-Sphingolipid desaturase from Brassica rapa

△~8-Sphingohpid desaturase is the key enzyme that catalyses desaturation at the C8 position of the long-chain base of sphingolipids in higher plants.There have been no previous studies on the genes encoding△~8-sphingolipid desaturases in Brassica rapa.In this study,four genes encoding A -sphingohpid desaturases from B.rapa were isolated and characterised.Phylogenetic analyses indicated that these genes could be divided into two groups:BrD8A,BrD8C and BrD8D in groupⅠ,and BrD8B in groupⅡ.The two groups of genes diverged before the separation of Arabidopsis and Brassica.Though the four genes shared a high sequence similarity,and their coding desaturases all located in endoplasmic reticulum,they exhibited distinct expression patterns.Heterologous expression in Saccharomyces cerevisiae revealed that BrD8A/B/C/D were functionally diverse A -sphingohpid desaturases that catalyse different ratios of the two products 8(Z)- and 8(E)-C18-phytosphingenine.The aluminium tolerance of transgenic yeasts expressing BrD8A/B/C/D was enhanced compared with that of control cells.Expression of BrD8A in Arabidopsis changed the ratio of 8(Z):8(E)-C18-phytosphingenine in transgenic plants. The information reported here provides new insights into the biochemical functional diversity and evolutionary relationship of△~8-sphingolipid desaturase in plants and lays a foundation for further investigation of the mechanism of 8(Z)- and 8(E)-C18-phytosphingenine biosynthesis.     

Isolation and identification of bovine tuberculosis in a Brazilian herd (São Paulo)

Mycobacterium was verified in animals from a Brazilian dairy herd, a total of 42 samples from 30 cows were submitted to culture and the isolated strains were analyzed by two polymerase chain reaction (PCR), the first specific for species belonging to the Mycobacterium complex (MTBC) and the other for differentiating M. tuberculosis from M. bovis. Twenty seven samples (64.3%) from 18 animals (60%) were positive for mycobacteria by culture, including samples from 15 retrofaryngeal lymphnodes (55.5%), 9 prescapular lymphnodes (33.3%), 2 lungs (7.4%), and 1 liver (3.7%). All isolated colonies were confirmed by PCR to contain MTBC organisms, and were identified as M. bovis by the same methodology.     

Isolation and structure elucidation of flavonoids from Amburana cearensis resin and identification of human DNA topoisomerase II-α inhibitors

Amburana cearensis, a plant in the Fabaceae family, is extensively used in folk medicine to treat grippes, coughs, bronchitis, sinusitis and external ulcers. Ultra-performance liquid chromatography coupled with diode array and quadrupole time-of-flight mass spectrometry (UPLC-DAD-QTOF-MS/MS) and combined use of data-independent acquisition (MSE) were used for profiling and structural characterization of the phenolic compounds observed in the resin of A. cearensis (16 isoflavones, 11 flavanonols, 3 chalcones, 1 isoflavonolignan and 1 isoflavoquinone). The isoflavonolignan (1”R,2”R-trans-1”-(7”-hydroxy-6”,8”-dimethoxyphenyl)-1′-(3′-hydroxy-4′-methoxyphenyl)-2”-hydroxymethyl-5-methoxy-1”,2”-dihydro-7H-1,4-dioxino[1”,2”-h]chromen-4-one) and an isoflavone (3′-hydroxy-7,8,4′-trimethoxyisoflavone) were identified as novel compounds. In this study, we isolated 15 principal phenolics and established their structures by different spectroscopic methods, including 1D and 2D NMR experiments, as well HR-ESI–MS analysis. The isolated reference compounds were used to explore fragmentation pathways. Compound identification was based on the exact mass, general fragmentation behaviors, retention times and UV absorption. The activity of the principal compounds against human DNA topoisomerase II-α was evaluated.     

Isolation and characterization of β-glucosidase from the cytosol of rat kidney cortex

A procedure is described for the preparation of extensively purified β-d-glucosidase (EC 3.2.1.21) from the cytosol fraction of rat kidney. The specific activity of the β-glucosidase in the high speed supernatant (100 000 × g, 90 min) fraction of rat kidney homogenate is 700-fold greater than that in the same fraction from heart, skeletal muscle, lung, spleen, brain or liver. β-Glucosidase activity co-chromatographs with β-d-galactosidase, β-d-fucosidase, α-l-arabinosidase and β-d-xylosidase activities through the last four column steps of the purification and their specific activities are 0.26, 0.39, 0.028 and 0.017 relative to that of β-glucosidase, respectively. The specific activity of the apparently homogeneous β-glucosidase is 115 000 nmol of glucose released from 4-methylumbelliferyl-β-d-glucopyranoside per mg protein per h. All five glycosidase activities possess similar pH dependency (pH optimum, 6–7) and heat lability, and co-migrate on polyacrylamide disc gels at ph 8.9 (R_F, 0.67). β-Glucosidase activity is inhibited competitively by glucono-(1 → 5)-lactone (K_I, 0.61 mM) and non-competitively by a variety of sulfhydryl reagents including N-ethylmaleimide, p-chloromercuribenzoate, 5,5′-dithio-bis(2-nitrobenzoic acid), and iodoacetic acid. Although the enzyme will release glucose from p-nitrophenyl and 4-methylumbelliferyl derivatives of β-d-glucose, it will not hydrolyze xylosyl-O-serine, β-d-glucocerebroside, lactose, galactosylovalbumin or trehalose. The enzyme consists of a single polypeptide chain with a molecular weight of 50 000–58 000, has a sedimentation coefficient of 4.41 S and contains a relatively large number of acidic amino acids. A study of the distribution of β-glucosidase activity in various regions of the dissected rat kidney indicates that the enzyme is probably contained in cells of the proximal convulated tubule. The enzyme is also present in relatively large ammounts in the villus cells, but not crypt cells, of the intestine. the physiological subtrates and function of the enzyme are unknown.     

Isolation, identification and quantitation of hydroxycinnamic acid conjugates, potential platform chemicals, in the leaves and stems of Miscanthus × giganteus using LC-ESI-MSn

Miscanthus × giganteus is a source of platform chemicals and bioethanol through fermentation. Cinnamates in leaves and stems were analysed by LC–ESI-MSⁿ. Free phenols were extracted and separated chromatographically. More than 20 hydroxycinnamates were identified by UV and LC–ESI-MSⁿ. Comparative LC–MS studies on the leaf extract showed isomers of O-caffeoylquinic acid (3-CQA, 4-CQA and 5-CQA), O-feruloylquinic acid (3-FQA, 4-FQA and 5-FQA) and para-coumaroylquinic acid (3-pCoQA and 5-pCoQA). Excepting 3-pCoQA, all were also detected in stem. 5-CQA dominated in leaf; a mandelonitrile–caffeoylquinic acid dominated in stem. Three minor leaf components were distinguished by fragmentation patterns in a targetted MS² experiment as dicaffeoylquinic acid isomers. Others (M_r 516) were tentatively identified as hexosylcaffeoyl-quinates. Three positional isomers of O-caffeoylshikimic acid were minor components. p-Hydroxybenzaldehyde was also a major component in stem. This is the first report of the hydroxycinnamic acid profile of leaves and stems of M. × giganteus.