Recent investigations show that exogenously applied small interfering RNAs (siRNA) and long double-stranded RNA (dsRNA) precursors can be taken up and translocated in plants to induce RNA interference (RNAi) in the plant or in its fungal pathogen. The question of whether genes in the plant genome can undergo suppression as a result of exogenous RNA application on plant surface is almost unexplored. This study analyzed whether it is possible to influence transcript levels of transgenes, as more prone sequences to silencing, in Arabidopsis genome by direct exogenous application of target long dsRNAs. The data revealed that in vitro synthesized dsRNAs designed to target the gene coding regions of enhanced green fluorescent protein (EGFP) or neomycin phosphotransferase II (NPTII) suppressed their transcript levels in Arabidopsis. The fact that, simple exogenous application of polynucleotides can affect mRNA levels of plant transgenes, opens new opportunities for the development of new scientific techniques and crop improvement strategies.

Exogenous application of double-stranded RNAs (dsRNAs) and small-interfering RNAs (siRNAs) to plant surfaces has emerged as a promising method for regulation of essential genes in plant pathogens and for plant disease protection. Yet, regulation of plant endogenous genes via external RNA treatments has not been sufficiently investigated. In this study, we targeted the genes of chalcone synthase (CHS), the key enzyme in the flavonoid/anthocyanin biosynthesis pathway, and two transcriptional factors, MYBL2 and ANAC032, negatively regulating anthocyanin biosynthesis in Arabidopsis. Direct foliar application of AtCHS-specific dsRNAs and siRNAs resulted in an efficient downregulation of the AtCHS gene and suppressed anthocyanin accumulation in A. thaliana under anthocyanin biosynthesis-modulating conditions. Targeting the AtMYBL2 and AtANAC032 genes by foliar dsRNA treatments markedly reduced their mRNA levels and led to a pronounced upregulation of the AtCHS gene. The content of anthocyanins was increased after treatment with AtMYBL2-dsRNA. Laser scanning microscopy showed a passage of Cy3-labeled AtCHS-dsRNA into the A. thaliana leaf vessels, leaf parenchyma cells, and stomata, indicating the dsRNA uptake and spreading into leaf tissues and plant individual cells. Together, these data show that exogenous dsRNAs were capable of downregulating Arabidopsis genes and induced relevant biochemical changes, which may have applications in plant biotechnology and gene functional studies.

The soft-shell clam Mya arenaria Linnaeus, 1758 is a commercially important fishery resource that occurs in boreal and temperate environments in the Northern Hemisphere. Whether the soft-shell clam is a single species with a circumboreal range or a species complex also comprising Mya japonica Jay, 1857 distributed in the north Pacific has long been debated by malacologists and palaeontologists based on slight differences in shell morphology. We used an integrative taxonomic approach incorporating available Mya spp. mitochondrial COI and 16S rRNA, and nuclear 28S rRNA gene sequences, as well as spermatozoan and shell morphological characters to test the validity of M. japonica and examine the range of soft-shell clam distribution. Although differences in shell morphology were minor, the results from tree topologies, pairwise uncorrected p-distances, Automatic Barcode Gap Discovery (ABGD) and spermatozoan ultramorphological data confirm the validity of M. japonica in both its endemic region in the northwest Pacific, and as here newly reported introduced populations in British Columbia in the northeast Pacific, and show that M. arenaria is distributed in the northeast Pacific, North Atlantic, Barents Sea (Arctic Ocean) and Mediterranean. We estimate these two closely related sister species diverged 4.1–12.5 Myr during early Pliocene to late Miocene, which is consistent with current evolutionary theory regarding M. arenaria. In addition, ABGD indicated the congener Mya truncata Linnaeus, 1758 may represent a species complex, but additional evidence is still needed to clarify its taxonomic status.

In bivalves neurotransmitters are involved in a variety of behaviors, but their diversity and distribution in the nervous system of these organisms remains somewhat unclear. Here, we first examined immunohistochemically the distributions of neurons containing different neurotransmitters, neuropeptides, and related enzymes, as well as the proliferative status of neurons in the ganglia of the mussel Crenomytilus grayanus. H-Phe-Met-Arg-Phe-NH2 (FMRFamide), choline acetyltransferase (ChAT), γ-aminobutyric acid (GABA) and tyrosine hydroxylase (TH) were found to be expressed by neurons in all the ganglia, whereas serotonin (5-HT) neurons were found only in the cerebropleural and pedal, but not visceral ganglia. Moreover, incubation of living mussels in the presence of a 5-HT precursor (5-HTP) confirmed the absence of 5-HT-containing neurons from the visceral ganglia, indicating that the “serotonin center” of the visceral nervous system is located in the cerebral ganglia. Furthermore, immunostaining of molecules related to neurotransmission together with α-acetylated tubulin demonstrated that this cytoskeletal protein may be a potential pan-neuronal marker in bivalves. Adult mussel neurons do not proliferate, but a population of proliferating PCNA-LIP cells which do not express any of the neurotransmitters examined, perhaps glia cells, was detected in the ganglia. These novel findings suggest that the nervous system of bivalves contains a broad variety of signal molecules most likely involved in the regulation of different physiological and behavioral processes. In addition, proliferating cells may maintain and renew glial cells and neurons throughout the lives of bivalves.

The present study focuses on two apparent species: the giant thyasirid Conchocele bisecta (Conrad, 1849), which is the dominant species of the benthic community in a gas hydrate area with cold-water methane-rich vents at a depth of about 800 m on the slope off Paramushir Island (Kuril Islands, Sea of Okhotsk) and small unidentified thyasirid bivalves from this same community. An examination of the shell morphology of these thyasirids showed that the small bivalves were in fact young specimens of C. bisecta, characterized by a high individual and age variability. A transmission electron microscopic study of C. bisecta revealed gills with ‘Type 3’ filaments, which were extended abfrontally and had a distinct bacteriocyte zone with extracellular symbionts. The symbiotic bacteria found were spherical, similar to thiotrophic symbionts of other thyasirids. The isotopic δ¹³С values of C. bisecta soft tissues (from −39.6 to −33.8‰) were much heavier than those of methane in the Paramushir gas-hydrate area and matched the range characteristic of symbiotrophic bivalves harbouring sulphur-oxidizing chemoautotrophic bacteria. The variations in δ¹³С and δ¹⁵N recorded for large and small C. bisecta can be related to ontogenetic differences in life habit: small individuals are totally buried in the sediment, while large ones are half-buried. Data from fatty acid (FA) analysis indicate that sulphur-oxidizing symbionts constitute almost the entire nutrition of C. bisecta, with no significant contribution of symbiotic or free-living methanotrophs. Furthermore, neither FA nor isotopic compositions provided evidence for photosynthetic sources as food items for C. bisecta through filter feeding.