Clam Worm Research Articles

DESS deconstructed: Is EDTA solely responsible for protection of high molecular weight DNA in this common tissue preservative?

DESS is a formulation widely used to preserve DNA in biological tissue samples. Although it contains three ingredients, dimethyl sulfoxide (DMSO), ethylenediaminetetraacetic acid (EDTA) and sodium chloride (NaCl), it is frequently referred to as a DMSO-based preservative. The effectiveness of DESS has been confirmed for a variety of taxa and tissues, however, to our knowledge, the contributions of each component of DESS to DNA preservation have not been evaluated. To address this question, we stored tissues of three aquatic taxa, Mytilus edulis (blue mussel), Faxonius virilis (virile crayfish) and Alitta virens (clam worm) in DESS, each component of DESS individually and solutions containing all combinations of two components of DESS. After storage at room temperature for intervals ranging from one day to six months, we extracted DNA from each tissue and measured the percentage of high molecular weight (HMW) DNA recovered (%R) and normalized HMW DNA yield (nY). Here, HMW DNA is defined as fragments >10 kb. For comparison, we also measured the %R and nY of HMW DNA from extracts of fresh tissues and those stored in 95% EtOH over the same time intervals. We found that in cases where DESS performed most effectively (yielding ≥ 20%R of HMW DNA), all solutions containing EDTA were as or more effective than DESS. Conversely, in cases where DESS performed more poorly, none of the six DESS-variant storage solutions provided better protection of HMW DNA than DESS. Moreover, for all taxa and storage intervals longer than one day, tissues stored in solutions containing DMSO alone, NaCl alone or DMSO and NaCl in combination resulted in %R and nY of HMW DNA significantly lower than those of fresh tissues. These results indicate that for the taxa, solutions and time intervals examined, only EDTA contributed directly to preservation of high molecular weight DNA.

Antioxidant and Anti-Inflammatory Effects of NCW Peptide from Clam Worm (Marphysa sanguinea).

Clam worms (Marphysa sanguinea) are a rich source of bioactive components such as the antibacterial peptide, perinerin. In the present study, we explored the physiological activities of a novel NCWPFQGVPLGFQAPP peptide (NCW peptide), which was purified from clam worm extract through high-performance liquid chromatography. Tandem mass spectrometry (MS/MS) revealed that NCW was a new peptide with a molecular weight of 1757.86 kDa. Moreover, NCW peptide exhibited significant antioxidant effects, causing a 50% inhibition of DPPH radical at a concentration of 20 μM without showing any cytotoxicity. These were associated with a reduction in the activity of catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and malondialdehyde (MDA) in LPS-stimulated RAW264. 7 cells. Furthermore, NCW peptide exhibited anti-inflammatory effects in LPS-stimulated RAW264.7 macrophages via inhibition of the abnormal production of pro-inflammatory cytokines including nitric oxide (NO), inducible nitric oxide synthase (iNOS), and cyclooxygenase-2 (COX-2). These anti-inflammatory effects of NCW peptide were associated with the inhibition of interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α). Our results therefore suggest that this novel NCW peptide with antioxidant and anti-inflammatory effects could be a good therapeutic agent against inflammation-related diseases.

Five polyketides isolated from the marine-derived fungus Arthrinium Sp.

Four new polyketides including two arthrinic acid derivatives (1–2), one phenolic derivative (3) and (S)-3-hydroxy-6-(2-hydroxypropyl)-5-methyl-2H-pyran-2-one (4) along with one methyl ester of arthrinic acid (5) were isolated from the culture broth of Arthrinium sp., which was an entophytic fungus of clam worm. Their structures were identified on the basis of HR-ESI-MS and NMR spectral analyses together with advanced Mosher's method. In the assay of inhibiting the prostate cancer PC3 cell line, none of the isolated compounds showed significant cytotoxicity.

Immunohistochemical localization of GnRH-immunoreactive cell bodies and fibers in the nerve ganglion of Perinereis aibuhitensis (Annelida: Polychaeta)

The gonadotropin-releasing hormone (GnRH) gene sequence has been identified in an annelid polychaete marine worm using continual genome sequencing. The distribution of GnRH immunoreactive (ir) cell bodies and fibers in the nerve ganglion of the clam worm Perinereis aibuhitensis (Polychaeta) was examined by immunohistochemistry using a newly produced rabbit polyclonal antibody raised against the marine worm GnRH (mwGnRH). The specificity of the antibody was confirmed by dot blot assay. The antibody cross-reacted with mwGnRH, but not with other forms of GnRH such as octopus GnRH, tunicate GnRH-I, II, owl limpet GnRH, and lamprey GnRH-II. In P. aibuhitensis, mwGnRH-ir cell bodies were detected in the nuclei 15–22, the caudal part of the cerebral ganglion. Furthermore, mwGnRH-ir fibers were mainly observed in the optic neuropil, but mwGnRH-ir fibers were also detected in the central neuropil region, the subpharyngeal ganglion, and the ventral nerve cord. These results indicate that mwGnRH is synthesized in the cerebral ganglion, is transported through the subpharyngeal ganglion and the ventral nerve cord, and functions either as a neurotransmitter or neuromodulator.

Population structuring and historical demography of a common clam worm Perinereris aibuhitensis near the coasts of Shandong Peninsula

Abstract To investigate the genetic diversity, population structure and historical demography of Perinereris aibuhitensis, a 684 base pair fragment of the mitochondrial DNA cytochrome oxidase subunit I gene was analyzed in 92 organisms sampled from around the coasts of Shandong Peninsula. The results showed low haplotype diversity and nucleotide diversity in P. aibuhitensis. Topologies of neighbor-joining trees were shallow but the frequencies of common haplotypes indicated recent evolutionary lineages in the associated geographic areas. Significant genetic differentiations were detected among the four sampled populations, reflecting limited gene flow. Neutrality tests and mismatch distributions showed population expansion in P. aibuhitensis, but a recent bottleneck or selection event is indicated for the samples in the Yellow River Estuary and the Jiaozhou Bay.

Glutathionyl-biliverdin IXα, a new heme catabolite in a marine annelid: Sex and cell specific accumulation

We have isolated and characterized the green pigment accumulating in coelomic cells (eleocytes) of the common clam worm Nereis virens by means of RP-HPLC and ESI-tandem mass spectrometry. This pigment is a novel biliverdin-glutathione conjugate in which the glutathione is linked to the biliverdin-backbone via a thioether bond. The yolk precursor vitellogenin, a female-specific high-density lipoprotein ( ρ = 1179 kg/m 3) with a native molecular mass of ∼500 kDa and a subunit mass of ∼150 kDa, is capable of transporting this pigment as well as heme. The sex-independent large discoidal lipoprotein present in the coelomic fluid, which is sequestered by the eleocytes, could also be shown to transport heme. This renders both lipoproteins as heme-lipoproteins. As the vitellogenin is secreted by the eleocytes, this suggests the eleocytes as a metabolic hub linking the uptake of the potent pro-oxidant heme thought to arise from aged hemoglobin via the large discoidal lipoprotein and its conversion to a bile pigment-conjugate. The conjugate is exported from the eleocytes to the oocytes via vitellogenin leading to the accumulation of green yolk protein. In contrast, no such export route exists in male eleocytes resulting in an accumulation of the biliverdin-conjugate in these cells.

Oyster reef community interactions: The effect of resident fauna on oyster ( Crassostrea spp.) larval recruitment

During their transition from the pelagic larval form to the benthic adult form, larvae are likely to encounter a diverse assemblage of resident invertebrates on oyster reefs. Fouling epifauna are generally believed to reduce the settlement of interspecific larvae through competitive exclusion and predation. Studies of these interactions, however, often utilize artificial settlement panels, which can exhibit different recruitment patterns to those observed on natural substrates. Native ( Crassostrea virginica) and non-native ( Crassostrea ariakensis) oyster larvae were exposed to reef-collected shells, each supporting a single species of reef-associated fauna, over a series of laboratory microcosm studies. The presence of adult bryozoans ( Membranipora tenuis) had little effect on rates of larval settlement or mortality. The boring sponge ( Cliona sp.) significantly decreased oyster larval settlement, and generally increased oyster mortality. Barnacles ( Balanus improvisus) typically facilitated settlement. Barnacle molds and empty barnacle tests, intended to mimic the surface area and rugosity of live barnacles, did not significantly affect settlement. In some trials, adult barnacle bathwater enhanced settlement of oyster larvae, highlighting the importance of waterborne cues. Barnacle bathwaters, adult clam worm ( Neanthes succinea) bathwaters and bathwaters created using adult oysters were all found to cause oyster larval mortality. Predation by clam worms, which were observed at very high densities on field-collected oyster shells, caused significant oyster larval mortality in these experiments. The combined effects of positive and negative interactions between oyster larvae and resident invertebrate reef fauna require enumeration under field conditions. The results from this study highlight the need for a more thorough understanding of settlement behavior and mortality sources of recruiting oyster larvae.

Mother Nature, Her Eels

1 Under streetlight, the line comes reeling in. It’s Friday night on the Seekonk River, and a Dominican immigrant has an American eel, Anguilla rostrata, on the hook. This is a “yellow eel,” slightly more than a foot long, thrashing in the shallows. More accurately, its color is green. It is the mature—but not yet sexually mature—stage of eel metamorphosis , one of many in the eel life cycle. This creature is likely three or four years old, and it’s probably a male. Females are larger, heavy with fecundity. Typically they are found higher upstream. It is apparent that this probable male isn’t going to make it. Ronny—a Guatemalan—comes to the aid of his Dominican friend, who is disinclined to deal with this thing, esta anguilla, wiggling maniacally. He grapples with it in the mud, locking a hand around the line, sliding it down over the slimy body. The body wraps around Ronny’s forearm and licks frightful circles, turning like a corkscrew. This eel, while rooting around in the benthic muck of this broad stretch of river, just swam upon a clam worm, una lumbriz, threaded with a hook. The worm was a heavy one, expensive too—twelve for nine dollars at a local bait shop on Branch Avenue. These fishermen pull them from a tangle of weeds in a white cardboard box, the kind that normally pops open to reveal pastries, but these pastries elongate as they are lifted, shaping themselves with round muscles, while nubby appendages finger down the sides of their bodies. Ronny had shown me in gestures how in Guatemala they take a machete and cut up the earth for similar long, legged worms. Now he grabs a rock with his right hand and takes it to the eel he has pinned between two stones. Blow after blow, he blasts it in the head. It is sudden and brutal, and I cringe. Yet the eel wriggles on—uncontrollably —seemingly unfazed by a broken skull. Ronny holds the body against his boot and takes his fingers to the snout, feeling for the remains of the lumbriz somewhere inside. The eel’s gape looks like an open wound; the creature looks like a snake sliced in half. Mother Nature, Her Eels Nick Neely 2 Ecotone: reimagining place “Fuck, mang,” says Ronny, “dat’s fucked up.” These are universal words, easy to appreciate. The eel squirms up and down in Ronny’s circled hand, defying grasp. I’d be cursing worse. “Man, it’s really down in there, isn’t it?” says Louis, a young Dominican, a driver for a company in Massachusetts. He is here with his mother, who is the girlfriend of the man whose line holds this eel. When I arrived, Louis was in the rear of a small sedan, smoking and drinking, hip-hop lyrics rolling out the window—not fishing. He spent most of his boyhood in Miami in “the hood,” as he says. Now he tours up and down the East Coast freighting perishables, turning west just three or four times a year to pick up or deliver. The barbed hook is lodged in flesh that’s come an awful long way only to be snared by a man living in Pawtucket, Rhode Island—all the way from the Sargasso Sea, where the eel was spawned. Eels are catadromous , returning from years spent in freshwater streams to breed and die in unknown depths near Bermuda. It’s a reverse paradigm to that of inland-born shad or salmon—anadromous fish. Eel courtship has never been documented, and we know little about their long journey. We do know that they don’t come back, that they mate and die. The genes of this one on the line won’t make it to the next generation; it’ll never return to the deep, calm water country where it was born. Ronny is the first of his family to live in the United States. His was a long journey of its own. He traveled at night, and in mass, like the eel. It took him seventeen days to trek on foot to the Texan border and cross it with...

Halogenated Veneers: Protein Cross‐Linking and Halogenation in the Jaws ofNereis, a Marine Polychaete Worm

Mineralized tissues are produced by most living organisms for load and impact functions. In contrast, the jaws of the clam worm, Nereis, are hard without mineralization. However, they are peculiarly rich in halogens, which are associated with a variety of post-translationally modified amino acids, many of which are multiply halogenated by chlorine, bromine, and/or iodine. Several of these modified amino acids, namely dibromohistidine, bromoiodohistidine, chloroiodotyrosine, bromoiodotyrosine, chlorodityrosine, chlorotrityrosine, chlorobromotrityrosine, and bromoiodotrityrosine, have not been previously reported. We have found that the distributions of Cl, Br, and I differ: Cl is widespread whereas Br and I, although not colocalized, are concentrated in proximity to the external jaw surfaces. By using nanoindentation, we show that Br and I are unlikely to play a purely mechanical role, but that the local Zn and Cl concentrations and jaw microstructure are the prime determinants of local jaw hardness. Several of the post-translationally modified amino acids are akin to those found in various sclerotized structures of invertebrates, and we propose that they are part of a cross-linked protein casing.

Angiotensin II and III upregulate body fluid volume of the clam worm Perinereis sp. via angiotensin II receptors in different manners

Angiotensin III (Ang III) as well as angiotensin II (Ang II) suppressed body weight loss of the clam worm Perinereis sp. under a hyper-osmotic condition, and enhanced body weight gain under a hypo-osmotic condition. Under a drying condition where the water inflow from outside the body was eliminated, Ang II suppressed body weight loss, but Ang III did not. Under these conditions, angiotensins I, IV, and (1–7) had no effect, and saralasin blocked the effects of Ang II and Ang III. It is concluded that Ang II and Ang III upregulate body fluid volume of the clam worm via Ang II receptors in different ways.

Angiotensin III as Well as Angiotensin II Regulates Water Flow through Aquaporins in a Clam Worm

Angiotensin III has been reported to exist in various animals and tissues. The physiological role, however, is still unclear except that brain angiotensin III is a central regulator of vasopressin release. In this study, angiotensin III as well as angiotensin II enhanced an increase in body weight of clam worms of Perinereis sp. under a hypo-osmotic condition and suppressed a decrease in body weight under a hyper-osmotic condition. When clam worms were treated with tetrachloroaurate (III) after angiotensin-treatment, these enhancing and suppressive effects of the angiotensins under hypo- and hyper-osmotic conditions were inhibited. In contrast, when clam worms were pretreated with tetrachloroaurate (III) before angiotensin-treatment, these effects of angiotensins were not inhibited. Since tetrachloroaurate (III) is a representative blocker of aquaporins, these results indicate that angiotensin III as well as angiotensin II regulates water flow through aquaporins in clam worms.

Evaluation of shrimp polyculture system in Thailand based on stable carbon and nitrogen isotope ratios

To quantify the contribution by cocultured animals to waste assimilation in an inten- sive shrimp farm in Thailand, the food web structures of the macrobenthos in a reservoir pond, a shrimp culture pond and water treatment ponds were examined using the stable C and N isotope ratio technique. Seawater for aquaculture was drawn from a creek, and stored in a reservoir pond, used for farming the banana prawn Fenneropenaeus merguiensis in culture ponds, and then recy- cled through treatment ponds where the green mussel Perna viridis was cultured to remove organic wastes discharged from the farming. The clam worm Nereididae sp. and the mud creeper Cerithideopsilla cingulata in the culture pond had d 13 C values of -21.0‰ and -18.4‰, respectively, suggesting that shrimp feed (mean d 13 C =- 20.7‰) was the main food source for these species. The d 13 C analysis also suggested that sediments (-23.7‰) in the reservoir pond and particulate organic matter (POM) (-24.0‰) and/or sediments (-25.0‰) in the treatment pond supplied carbon for most macrobenthic animals. However, green mussels in the treatment pond had a mean d 13 C value of -20.5‰, suggesting that shrimp feed was the main food source for this species.

Two 1-Alkyl-2-acyl Choline Glycerophospholipids Having an Arachidonoyl or Eicosapentaenoyl Group, from the Clam Worm (Marphysa sanguinea).

Ten choline glucerophospholipids (CGP) were isolated from the marine annelid, Marphysa sanguinea. Among them, six compounds were lyso platelet-activating factors (PAFs) and two were 1-hexadecyl-2-arachidonoyl-and 1-hexadecyl-2-eicosapentanoyl-sn-glycero-3-phosphocholines, which are regarded as stored precursor forms of PAF, so-called PAF-like lipid. These two compounds comprised 28% of the alkylacyl CGP fraction and are considered to be the main components of PAF-like lipid in this material.

Phosphocholine-Bonded Galactosylceramides Having a Tri-Unsaturated Long-Chain Base from the Clam Worm, Marphysa sanguinea

Seven glycosphingolipids were obtained in the pure state from the clam worm, Marphysa sanguinea, by preparative HPLC with a reversed-phase column in a recycling mode. Their structures were elucidated based on detailed analyses of 1H- and 13C-NMR spectra of the intact compounds. The position and the geometry of double bonds in the long-chain base (LCB) were determined by the two-dimensional NMR (COSY, NOESY, and HMBC) analyses. All compounds are phosphocholine-bonded monogalactosylceramides and two of them have tri-unsaturated long-chain bases.

Distribution of alkyl and alkenyl ether-linked phospholipids and platelet-activating factor-like lipid in various species of invertebrates

The levels of alkenylacyl, alkylacyl and diacyl subclasses of choline glycerophospholipid (CGP) and ethanolamine glycerophospholipid (EGP) fractions in 28 species of various invertebrates were studied. We found that only small amounts of either 1-alkyl-2acyl- sn-phosphocholine (alkylacyl-GPC) or 1-alkenyl-2-acyl- sn-glycero-3-phosphoethanolamine (alkenylacyl-GPE) are present in most species of insects. On the other hand, almost all species examined in various phyla other than Arthropoda were shown to contain large amounts of both alkylacyl-GPC and alkenylacyl-GPE. The highest proportion of alkylacyl subclass in CGP was noted in sponge, Halichondria japonica (81.8% of CGP) and the highest proportion of alkenylacyl subclass in EGP was found in clam worm, Marphysa sanguinea (88.7% of EGP). We next surveyed the presence of platelet-activating factor (PAF)-like lipid in 45 species of invertebrates. PAF-like lipid was widely distributed among various lower animals. The highest value was obtained for sea cucumber, Stichopus japonicas, in which PAF-like lipid was present throughout the body. We also confirmed the presence of acetyltransferase activity in several lower animals. These results suggest that alkyl and alkenyl ether-linked phospholipids including PAF are physiologically important molecules particularly for invertebrates belonging to lower phyla.

Hetero‐α‐helical, two‐chain coiled‐coils. Clam–worm hybrid paramyosins

AbstractThe specificity of the interaction between the α‐helices in two‐chain coiled‐coils is investigated by studying the formation of hybrid molecules in which one α‐helix is a clam paramyosin chain and the other a worm paramyosin chain. Hybrids are formed by mixing, denaturation, and subsequent renaturation. Comparison is made with a blank solution in which renaturation precedes mixing, thus precluding hybridization. Hybrids are detected by a ruse based on the presence of free sulfhydryl functions on calm chains. This allows molecules comprising two clam chains to be covalently crosslinked by oxidation with 5,5′‐dithiobis(2‐nitrobenzoate). Worm paramyosin chains have no sulfhydryl, so molecules comprising two worm chains or hybrid molecules comprising one chain of each type cannot crosslink. When run on sodium dodecyl sulfate polyacrylamide gel electrophoresis, therefore, the protein separates into two well‐resolved regions, one containing one‐chain species and the other two‐chain species. When the gels are scanned and quantitated, the hybrids show up as an increase in the fraction of material in the one‐chain band compared with the fraction in the blank solution. When renaturation is direct, we find that the fraction of renaturated molecules that are hybrids varies from ∼10% at 5°C to ∼5% at 25°C. These are judged to be nonequilibrium (quenched) values. When renaturation is by slow annealing, the equilibrium fraction hybrids are ∼4% and show a modest, but measurable, increase with increasing temperature. These data allow calculation of the equilibrium constant Kh and standard free energy for the hybridization reaction: (1/2)CC + (½)WW = CW, in which C(W) stands for an α‐helical clam (worm) polypeptide chain. The temperature dependence gives the standard enthalpy and entropy of the reaction. We find ΔH ≃ 1800 cal mol−1 and ΔH ≃ 1.4 cal mol−1 K−1, using molarity concentration units and the infinitely dilute solution in NaCl/phosphate buffer as reference state. The possible molecular significance of these values is discussed, and it is concluded that the observed standard entropy arises essentially entirely from the rotational dissymmetry of the hybrids.

OBSERVATIONS ON CERCARIÆUM LINTONI MILLER AND NORTHUP AND ITS METACERCARIAL DEVELOPMENT

The description of Cercariæum lintoni Miller and Northup as given by earlier workers is supplemented and emended in the light of knowledge obtained by comparing the cercariæum with its metacercarial stage, developed in experimental animals. The true nature of the digestive and reproductive systems is made clear.Cercariæum lintoni penetrated the parapodia of Nereis virens, encysted, and developed into the metacercarial stage. Although the cercaria would penetrate and encyst in other invertebrates, the clam worm was the only form in which development was secured. Nereis virens appears to be a natural host. The metacercaria develops adult features as the larval characters disappear.It is difficult to determine the relationship of this species to other trematode groups; the digestive system is similar to that of the species of the family Haploporidæ, but the arrangement of the reproductive organs is very different and resembles that of the Zoogonidæ. palombi's suggestions concerning the relations of the species are discussed.