Interplay between Two Bacterial Actin Homologs,MamK and MamK-Like,Is Required for the Alignment of Magnetosome Organelles in Magnetospirillum magneticum AMB-1

Many bacterial species contain multiple actin-like proteins tasked with the execution of crucial cell biological functions. MamK, an actin-like protein found in magnetotactic bacteria, is important in organizing magnetosome organelles into chains that are used for navigation along geomagnetic fields. MamK and numerous other magnetosome formation factors are encoded by a genetic island termed the magnetosome island. Unlike most magnetotactic bacteria, Magnetospirillum magneticum AMB-1 (AMB-1) contains a second island of magnetosome-related genes that was named the magnetosome islet. A homologous copy of mamK, mamK-like, resides within this islet and encodes a protein capable of filament formation in vitro. Previous work had shown that mamK-like is expressed in vivo, but its function, if any, had remained unknown. Though MamK-like is highly similar to MamK, it contains a mutation that in MamK and other actins blocks ATPase activity in vitro and filament dynamics in vivo. Here, using genetic analysis, we demonstrate that mamK-like has an in vivo role in assisting organelle alignment. In addition, MamK-like forms filaments in vivo in a manner that is dependent on the presence of MamK and the two proteins interact in a yeast two-hybrid assay. Surprisingly, despite the ATPase active-site mutation, MamK-like is capable of ATP hydrolysis in vitro and promotes MamK filament turnover in vivo. Taken together, these experiments suggest that direct interactions between MamK and MamK-like contribute to magnetosome alignment in AMB-1.     

Screening for the Interacting Partners of the Proteins MamK & MamJ by Two-Hybrid Genomic DNA Library of Magnetospirillum magneticum AMB-1

Magnetotactic bacteria are a group of prokaryotes capable of sensing and navigating along the earth’s magnetic field. The linear alignment of magnetosomes, which acts as a compass needle for orientation, is dependent on the proteins MamJ (amb0964) & MamK (amb0965). We constructed Magnetospirillum magneticum AMB-1 two-hybrid DNA libraries by fusing the random genomic fragments of AMB-1 to the N-terminal domain of the α-subunit of RNA polymerase in vector pTRG and used as preys. The genes mamJ & mamK were cloned in frame with the λ repressor protein (λ cI) in vector pBT and used as baits for screening the binding partners. After preliminary screening, we further confirmed the candidate interactions between selected protein pairs. The results showed that there were relatively strong interactions between MamK versus Amb3498 (flagella motor switch protein fliM), versus Amb0854 MCPs (signal domain of methyl-accepting chemotaxis protein) and versus Amb3568 (GGDEF domain-containing protein), respectively. MamJ versus Amb1722 (hypothetical protein), MamJ versus MamK, and MamK versus Amb1807 (cation transport ATPase) exhibited low level of interaction. Although the TPR repeat protein MamA (amb0971) showed no interaction with either MamJ or MamK, the TPR repeat protein Amb0024 with more motif sequences exhibited relatively strong interaction with MamK. Among the identified proteins, all categorized as signal transduction-related displayed interaction only with MamK and without MamJ, suggesting that magnetotaxis via MamK in Magnetospirillum magneticum AMB-1 might be somehow concerned with the widely accepted chemotaxis mechanism in bacteria.     

An MCP-Like Protein Interacts with the MamK Cytoskeleton and Is Involved in Magnetotaxis in Magnetospirillum magneticum AMB-1

Magnetotactic bacteria have the unique capacity of aligning and swimming along geomagnetic field lines, a behavior called magnetotaxis. Although this behavior has been observed for 40 years, little is known about its mechanism. Magnetotactic bacteria synthesize unique organelles, magnetosomes, which are magnetic crystals enveloped by membrane. They form chains with the help of the filamentous cytoskeletal protein MamK and impart a net magnetic-dipole moment to the bacterium. The current model proposes that magnetotaxis comprises passive magnetic orientation and active swimming due to flagellar rotation. We thought that magnetic sensing, via the widely used chemotaxis mechanism, might be actively involved in magnetotaxis. We found that the methyl-accepting chemotaxis protein Amb0994 of Magnetospirillum magneticum AMB-1 was capable of carrying out such a function. Amb0994 is encoded by a gene in the magnetosome island, in which genes essential for magnetosome biosynthesis and magnetotaxis are concentrated. Amb0994 lacks periplasmic sensing domain, which is generally involved in sensing stimuli from outside of cells. By constructing fusions with a derivative of yellow-fluorescent-protein, we showed that Amb0994 localizes to the cell poles, where methyl-accepting chemotaxis proteins are usually clustered. We then showed that Amb0994 specifically interacts, via its C-terminal domain, with MamK, using a bimolecular fluorescence complementation assay. Moreover, overproduction of Amb0994 slowed down the response of the bacterium to changes in the direction of the magnetic field. Most importantly, the C-terminal domain of Amb0994, which interacts with MamK, is responsible for this phenotype, suggesting that the interaction between Amb0994 and MamK plays a key role in magnetotaxis. These results lead to a novel explanation for magnetotaxis at the molecular level.     

Insight into the assembly properties and functional organisation of the magnetotactic bacterial actin-like homolog, MamK

Magnetotactic bacteria (MTB) synthesize magnetosomes, which are intracellular vesicles comprising a magnetic particle. A series of magnetosomes arrange themselves in chains to form a magnetic dipole that enables the cell to orient itself along the Earth's magnetic field. MamK, an actin-like homolog of MreB has been identified as a central component in this organisation. Gene deletion, fluorescence microscopy and in vitro studies have yielded mechanistic differences in the filament assembly of MamK with other bacterial cytoskeletal proteins within the cell. With little or no information on the structural and behavioural characteristics of MamK outside the cell, the mamK gene from Magnetospirillium gryphiswaldense was cloned and expressed to better understand the differences in the cytoskeletal properties with its bacterial homologues MreB and acitin. Despite the low sequence identity shared between MamK and MreB (22%) and actin (18%), the behaviour of MamK monitored by light scattering broadly mirrored that of its bacterial cousin MreB primarily in terms of its pH, salt, divalent metal-ion and temperature dependency. The broad size variability of MamK filaments revealed by light scattering studies was supported by transmission electron microscopy (TEM) imaging. Filament morphology however, indicated that MamK conformed to linearly orientated filaments that appeared to be distinctly dissimilar compared to MreB suggesting functional differences between these homologues. The presence of a nucleotide binding domain common to actin-like proteins was demonstrated by its ability to function both as an ATPase and GTPase. Circular dichroism and structural homology modelling showed that MamK adopts a protein fold that is consistent with the 'classical' actin family architecture but with notable structural differences within the smaller domains, the active site region and the overall surface electrostatic potential.     

A Second Actin-Like MamK Protein in Magnetospirillum magneticum AMB-1 Encoded Outside the Genomic Magnetosome Island

Magnetotactic bacteria are able to swim navigating along geomagnetic field lines. They synthesize ferromagnetic nanocrystals that are embedded in cytoplasmic membrane invaginations forming magnetosomes. Regularly aligned in the cytoplasm along cytoskeleton filaments, the magnetosome chain effectively forms a compass needle bestowing on bacteria their magnetotactic behaviour. A large genomic island, conserved among magnetotactic bacteria, contains the genes potentially involved in magnetosome formation. One of the genes, mamK has been described as encoding a prokaryotic actin-like protein which when it polymerizes forms in the cytoplasm filamentous structures that provide the scaffold for magnetosome alignment. Here, we have identified a series of genes highly similar to the mam genes in the genome of Magnetospirillum magneticum AMB-1. The newly annotated genes are clustered in a genomic islet distinct and distant from the known magnetosome genomic island and most probably acquired by lateral gene transfer rather than duplication. We focused on a mamK-like gene whose product shares 54.5% identity with the actin-like MamK. Filament bundles of polymerized MamK-like protein were observed in vitro with electron microscopy and in vivo in E. coli cells expressing MamK-like-Venus fusions by fluorescence microscopy. In addition, we demonstrate that mamK-like is transcribed in AMB-1 wild-type and ΔmamK mutant cells and that the actin-like filamentous structures observed in the ΔmamK strain are probably MamK-like polymers. Thus MamK-like is a new member of the prokaryotic actin-like family. This is the first evidence of a functional mam gene encoded outside the magnetosome genomic island.     

The FtsZ-Like Protein FtsZm of Magnetospirillum gryphiswaldense Likely Interacts with Its Generic Homolog and Is Required for Biomineralization under Nitrate Deprivation

Midcell selection, septum formation, and cytokinesis in most bacteria are orchestrated by the eukaryotic tubulin homolog FtsZ. The alphaproteobacterium Magnetospirillum gryphiswaldense (MSR-1) septates asymmetrically, and cytokinesis is linked to splitting and segregation of an intracellular chain of membrane-enveloped magnetite crystals (magnetosomes). In addition to a generic, full-length ftsZ gene, MSR-1 contains a truncated ftsZ homolog (ftsZm) which is located adjacent to genes controlling biomineralization and magnetosome chain formation. We analyzed the role of FtsZm in cell division and biomineralization together with the full-length MSR-1 FtsZ protein. Our results indicate that loss of FtsZm has a strong effect on microoxic magnetite biomineralization which, however, could be rescued by the presence of nitrate in the medium. Fluorescence microscopy revealed that FtsZm-mCherry does not colocalize with the magnetosome-related proteins MamC and MamK but is confined to asymmetric spots at midcell and at the cell pole, coinciding with the FtsZ protein position. In Escherichia coli, both FtsZ homologs form distinct structures but colocalize when coexpressed, suggesting an FtsZ-dependent recruitment of FtsZm. In vitro analyses indicate that FtsZm is able to interact with the FtsZ protein. Together, our data suggest that FtsZm shares key features with its full-length homolog but is involved in redox control for magnetite crystallization.     

In vitro assembly of the bacterial actin protein MamK from ‘Candidatus Magnetobacterium casensis’ in the phylum Nitrospirae

Magnetotactic bacteria (MTB), a group of phylogenetically diverse organisms that use their unique intracellular magnetosome organelles to swim along the Earth’s magnetic field, play important roles in the biogeochemical cycles of iron and sulfur. Previous studies have revealed that the bacterial actin protein MamK plays essential roles in the linear arrangement of magnetosomes in MTB cells belonging to the Proteobacteria phylum. However, the molecular mechanisms of multiple- magnetosome-chain arrangements in MTB remain largely unknown. Here, we report that the MamK filaments from the uncultivated ‘Candidatus Magnetobacterium casensis’ (Mcas) within the phylum Nitrospirae polymerized in the presence of ATP alone and were stable without obvious ATP hydrolysis-mediated disassembly. MamK in Mcas can convert NTP to NDP and NDP to NMP, showing the highest preference to ATP. Unlike its Magnetospirillum counterparts, which form a single magnetosome chain, or other bacterial actins such as MreB and ParM, the polymerized MamK from Mcas is independent of metal ions and nucleotides except for ATP, and is assembled into well-ordered filamentous bundles consisted of multiple filaments. Our results suggest a dynamically stable assembly of MamK from the uncultivated Nitrospirae MTB that synthesizes multiple magnetosome chains per cell. These findings further improve the current knowledge of biomineralization and organelle biogenesis in prokaryotic systems.     

MamK, a bacterial actin, forms dynamic filaments in vivo that are regulated by the acidic proteins MamJ and LimJ

Bacterial actins, in contrast to their eukaryotic counterparts, are highly divergent proteins whose wide-ranging functions are thought to correlate with their evolutionary diversity. One clade, represented by the MamK protein of magnetotactic bacteria, is required for the subcellular organization of magnetosomes, membrane-bound organelles that aid in navigation along the earth's magnetic field. Using a fluorescence recovery after photobleaching assay in Magnetospirillum magneticum AMB-1, we find that, like traditional actins, MamK forms dynamic filaments that require an intact NTPase motif for their turnover in vivo. We also uncover two proteins, MamJ and LimJ, which perform a redundant function to promote the dynamic behaviour of MamK filaments in wild-type cells. The absence of both MamJ and LimJ leads to static filaments, a disrupted magnetosome chain, and an anomalous build-up of cytoskeletal filaments between magnetosomes. Our results suggest that MamK filaments, like eukaryotic actins, are intrinsically stable and rely on regulators for their dynamic behaviour, a feature that stands in contrast to some classes of bacterial actins characterized to date.     

The acidic repetitive domain of the Magnetospirillum gryphiswaldense MamJ protein displays hypervariability but is not required for magnetosome chain assembly

Magnetotactic bacteria navigate along the earth's magnetic field using chains of magnetosomes, which are intracellular organelles comprising membrane-enclosed magnetite crystals. The assembly of highly ordered magnetosome chains is under genetic control and involves several specific proteins. Based on genetic and cryo-electron tomography studies, a model was recently proposed in which the acidic MamJ magnetosome protein attaches magnetosome vesicles to the actin-like cytoskeletal filament formed by MamK, thereby preventing magnetosome chains from collapsing. However, the exact functions as well as the mode of interaction between MamK and MamJ are unknown. Here, we demonstrate that several functional MamJ variants from Magnetospirillum gryphiswaldense and other magnetotactic bacteria share an acidic and repetitive central domain, which displays an unusual intra- and interspecies sequence polymorphism, probably caused by homologous recombination between identical copies of Glu- and Pro-rich repeats. Surprisingly, mamJ mutant alleles in which the central domain was deleted retained their potential to restore chain formation in a DeltamamJ mutant, suggesting that the acidic domain is not essential for MamJ's function. Results of two-hybrid experiments indicate that MamJ physically interacts with MamK, and two distinct sequence regions within MamJ were shown to be involved in binding to MamK. Mutant variants of MamJ lacking either of the binding domains were unable to functionally complement the DeltamamJ mutant. In addition, two-hybrid experiments suggest both MamK-binding domains of MamJ confer oligomerization of MamJ. In summary, our data reveal domains required for the functions of the MamJ protein in chain assembly and maintenance and provide the first experimental indications for a direct interaction between MamJ and the cytoskeletal filament protein MamK.     

How magnetotactic bacteria make magnetosomes queue up

Magnetotactic bacteria contain chains of magnetosomes that comprise a permanent magnetic dipole in each cell. In two separate, recent papers, Scheffel et al. and Komeili et al. describe the roles of the proteins MamJ and MamK in magnetosome chain formation. Here, we describe the two studies and highlight questions that must be addressed in future investigations of how magnetotactic bacteria construct their magnetic compass needles.     

Proteomic analysis of irregular,bullet‐shaped magnetosomes in the sulphate‐reducing magnetotactic bacterium Desulfovibrio magneticus RS‐1

Recent molecular studies on magnetotactic bacteria have identified a number of proteins associated with bacterial magnetites (magnetosomes) and elucidated their importance in magnetite biomineralisation. However, these analyses were limited to magnetotactic bacterial strains belonging to the α‐subclass of Proteobacteria. We performed a proteomic analysis of magnetosome membrane proteins in Desulfovibrio magneticus strain RS‐1, which is phylogenetically classified as a member of the δ‐Proteobacteria. In the analysis, the identified proteins were classified based on their putative functions and compared with the proteins from the other magnetotactic bacteria, Magnetospirillum magneticum AMB‐1 and M. gryphiswaldense MSR‐1. Three magnetosome‐specific proteins, MamA (Mms24), MamK, and MamM, were identified in strains RS‐1, AMB‐1, and MSR‐1. Furthermore, genes encoding ten magnetosome membrane proteins, including novel proteins, were assigned to a putative magnetosome island that contains subsets of genes essential for magnetosome formation. The collagen‐like protein and putative iron‐binding proteins, which are considered to play key roles in magnetite crystal formation, were identified as specific proteins in strain RS‐1. Furthermore, genes encoding two homologous proteins of Magnetococcus MC‐1 were assigned to a cryptic plasmid of strain RS‐1. The newly identified magnetosome membrane proteins might contribute to the formation of the unique irregular, bullet‐shaped crystals in this microorganism.     

Polar positional information in Escherichia coli spherical cells

Shigella surface protein IcsA and its cytoplasmic derivatives are localized to the old pole of rod-shaped cells when expressed in Escherichia coli. In spherical mreB cells, IcsA is targeted to ectopic sites and close to one extremity of actin-like MamK filament. To gain insight into the properties of the sites containing polar material, we studied the IcsA localization in spherical cells. GFP was exported into the periplasm via the Tat pathway and used as a periplasmic space marker. GFP displayed zonal fluorescence in both mreB and rodA-pbpA spherical E. coli cells, indicating an uneven periplasmic space. Deconvolution images revealed that the cytoplasmic IcsA fused to mCherry was localized outside or at the edge of the GFP zones. These observations strongly suggest that polar material is restricted to the positions where the periplasm possesses particular structural or biochemical properties.     

细菌纳米磁小体的合成机制及应用

综述了近年趋磁细菌纳米磁小体生物合成的分子机制及应用进展。磁小体的合成涉及磁小体膜的形成、铁的吸收和转运、磁小体晶体的矿化、成熟以及磁小体的链状排列等。其中Mam J和Mam K互作并丝状排列,固定磁小体使其链状排列及磁小体膜由细胞质膜内陷而形成是两个令人注目的成就。我们也提出了关于磁小体的生理意义及合成机制的假说:细胞在低氧浓度下由于氧胁迫大量吸收铁,Fe³⁺/Fe²⁺电子对可起到类似O₂/H₂O的作用,产生能量并作为电子受体;Fe3+得到电子还原成的Fe²⁺可引起Fenton反应,此反应产生的活性氧可影响到生物体的正常生理代谢,细胞为降低Fe²⁺浓度,将其与Fe³⁺一同转化为Fe₃O₄颗粒;磁小体的生理功能之一是降低胞内的活性氧。     

A new species of Hemibrycon (Characiformes,Characidae) from the upper San Juan River drainage,Pacific versant,Colombia

Hemibrycon sanjuanensis, new species, is described from the upper San Juan River drainage, Pacific versant, Colombia. It is distinguished from Hemibrycon boquiae, Hemibrycon brevispini, Hemibrycon cairoense, Hemibrycon colombianus, Hemibrycon mikrostiktos, Hemibrycon metae, Hemibrycon palomae, Hemibrycon rafaelense and Hemibrycon tridens by the presence of a circular or oblong humeral spot that is located two scales posterior to the opercle (vs. 3–4 scales in Hemibrycon palomae, Hemibrycon rafaelense, Hemibrycon brevispini and Hemibrycon cairoense, and 0–1 scales, in Hemibrycon metae and Hemibrycon boquiae). It further differs from Hemibrycon colombianus in having a round or oblong humeral spot (vs. rectangular). It differs from Hemibrycon beni, Hemibrycon dariensis, Hemibrycon divisorensis, Hemibrycon helleri, Hemibrycon huambonicus, Hemibrycon inambari, Hemibrycon jabonero, Hemibrycon jelskii, Hemibrycon mikrostiktos, Hemibrycon polyodon, Hemibrycon quindos, Hemibrycon raqueliae, Hemibrycon santamartae, Hemibrycon surinamensis, Hemibrycon taeniurus, Hemibrycon tridens, and Hemibrycon yacopiae in having melanophores on the posterior margins of the scales along the sides of body (vs. lacking melanophores on margins of scales along entire length of the sides of body). The new species differs from all congeners mentioned above in having, among other features, six teeth in the outer premaxillary row arranged in a straight line (vs. five or fewer teeth not arranged in straight line except Hemibrycon cairoense with two to six teeth in the outer premaxillary row).     

Salt lakes of La Mancha (Central Spain): A hot spot for tiger beetle (Carabidae,Cicindelinae) species diversity

The tiger beetle assemblage of the wetlands of La Mancha (central Spain) comprises nine species: Calomera littoralis littoralis, Cephalota maura maura, Cephalota circumdata imperialis, Cephalota dulcinea, Cicindela campestris campestris, Cicindela maroccana, Cylindera paludosa, Lophyra flexuosa flexuosa, and Myriochila melancholica melancholica. This assemblage represents the largest concentration of tiger beetles in a single 1º latitude / longitude square in Europe. General patterns of spatial and temporal segregation among species are discussed based on observations of 1462 specimens registered during an observation period of one year, from April to August. The different species of Cicindelini appear to be distributed over space and time, with little overlapping among them. Three sets of species replace each other phenologically as the season goes on. Most of the species occupy drying or dried salt lakes and salt marshes, with sparse vegetation cover. Spatial segregation is marked in terms of substrate and vegetation use. Calomera littoralis and Myriochila melancholica have been observed mainly on wet soils; Cephalota circumdata on dry open saline flats; Cephalota dulcinea and Cylindera paludosa in granulated substrates with typical halophytic vegetation; Cephalota maura is often present in man-modified areas. Cephalota circumdata and Cephalota dulcinea are included as species of special interest in the list of protected species in Castilla–La Mancha. Conservation problems for the Cicindelini assemblage arise from agricultural activities and inadequate use of sport vehicles. Attempts at restoring the original habitat, supressing old semi-industrial structures, may affect the spatial heterogeneity of the lakes, and have an effect on Cicindelinae diversity.     

Taxonomic hypotheses regarding the genus Gerbillus (Rodentia,Muridae, Gerbillinae) based on molecular analyses of museum specimens

Methodological improvements now allow routine analyses of highly degraded DNA samples as found in museum specimens. Using these methods could be useful in studying such groups as rodents of the genus Gerbillus for which i) the taxonomy is still highly debated, ii) collection of fresh specimens may prove difficult. Here we address precise taxonomic questions using a small portion of the cytochrome b gene obtained from 45 dry skin/skull museum samples (from 1913 to 1974) originating from two African and three Asian countries. The specimens were labelled Gerbillus gerbillus, Gerbillus andersoni, Gerbillus nanus, Gerbillus amoenus, Gerbillus perpallidus and Gerbillus pyramidum, and molecular results mostly confirmed these assignations. The close relationship between Gerbillus nanus (Asian origin) and Gerbillus amoenus (African origin) confirmed that they represent vicariant sibling species which differentiated in allopatry on either side of the Red Sea. In the closely related Gerbillus perpallidus and Gerbillus pyramidum, specimens considered as belonging to one Gerbillus pyramidum subspecies (Gerbillus pyramidum floweri) appeared closer to Gerbillus perpallidus suggesting that they (Gerbillus pyramidum floweri and Gerbillus perpallidus) may represent a unique species, distributed on both sides of the Nile River, for which the correct name should be Gerbillus floweri. Furthermore, the three other Gerbillus pyramidum subspecies grouped together with no apparent genetic structure suggesting that they may not yet represent genetically differentiated lineages. This study confirms the importance of using these methods on museum samples, which can open new perspectives in this particular group as well as in other groups of interest.     

An Extensive Comparison of the Effect of Anthelmintic Classes on Diverse Nematodes

Soil-transmitted helminths are parasitic nematodes that inhabit the human intestine. These parasites, which include two hookworm species, Ancylostoma duodenale and Necator americanus, the whipworm Trichuris trichiura , and the large roundworm Ascaris lumbricoides , infect upwards of two billion people and are a major cause of disease burden in children and pregnant women. The challenge with treating these diseases is that poverty, safety, and inefficient public health policy have marginalized drug development and distribution to control infection in humans. Anthelmintics (anti-worm drugs) have historically been developed and tested for treatment of non-human parasitic nematodes that infect livestock and companion animals. Here we systematically compare the in vitro efficacy of all major anthelmintic classes currently used in human therapy (benzimidazoles, nicotinic acetylcholine receptor agonists, macrocyclic lactones, nitazoxanide) against species closely related to human parasitic nematodes-Ancylostoma ceylanicum, Trichuris muris , and Ascaris suum --- as well as a rodent parasitic nematode used in veterinary drug discovery, Heligmosomoides bakeri , and the free-living nematode Caenorhabditis elegans. Extensive in vitro data is complemented with single-dose in vivo data in three rodent models of parasitic diseases. We find that the effects of the drugs in vitro and in vivo can vary greatly among these nematode species, e.g., the efficacy of albendazole is strong on A. ceylanicum but weak on H . bakeri . Nonetheless, certain commonalities of the in vitro effects of the drugs can be seen, e.g., nitazoxanide consistently shows an all-or-nothing response. Our in vitro data suggest that further optimization of the clinical efficacy of some of these anthelmintics could be achieved by altering the treatment routine and/or dosing. Most importantly, our in vitro and in vivo data indicate that the hookworm A. ceylanicum is a particularly sensitive and useful model for anthelmintic studies and should be incorporated early on in drug screens for broad-spectrum human soil-transmitted helminth therapies.     

The Complete Plastid Genome Sequence of Madagascar Periwinkle Catharanthus roseus (L.) G. Don: Plastid Genome Evolution,Molecular Marker Identification,and Phylogenetic Implications in Asterids

The Madagascar periwinkle ( Catharanthus roseus in the family Apocynaceae) is an important medicinal plant and is the source of several widely marketed chemotherapeutic drugs. It is also commonly grown for its ornamental values and, due to ease of infection and distinctiveness of symptoms, is often used as the host for studies on phytoplasmas, an important group of uncultivated plant pathogens. To gain insights into the characteristics of apocynaceous plastid genomes (plastomes), we used a reference-assisted approach to assemble the complete plastome of C . roseus , which could be applied to other C . roseus -related studies. The C . roseus plastome is the second completely sequenced plastome in the asterid order Gentianales. We performed comparative analyses with two other representative sequences in the same order, including the complete plastome of Coffea arabica (from the basal Gentianales family Rubiaceae) and the nearly complete plastome of Asclepias syriaca (Apocynaceae). The results demonstrated considerable variations in gene content and plastome organization within Apocynaceae, including the presence/absence of three essential genes (i.e., accD, clpP, and ycf1) and large size changes in non-coding regions (e.g., rps2-rpoC2 and IRb-ndhF). To find plastome markers of potential utility for Catharanthus breeding and phylogenetic analyses, we identified 41 C . roseus -specific simple sequence repeats. Furthermore, five intergenic regions with high divergence between C . roseus and three other euasterids I taxa were identified as candidate markers. To resolve the euasterids I interordinal relationships, 82 plastome genes were used for phylogenetic inference. With the addition of representatives from Apocynaceae and sampling of most other asterid orders, a sister relationship between Gentianales and Solanales is supported.     

Further contributions to the longhorn beetle (Coleoptera,Cerambycidae) fauna of New Brunswick and Nova Scotia,Canada

Sixteen species of Cerambycidae are newly recorded for New Brunswick, Canada; Arhopalus obsoletus (Randall), Atimia confusa confusa (Say), Callidium frigidum Casey, Phymatodes amoenus (Say), Phymatodes testaceus (Linnaeus), Neoclytus mucronatus mucronatus (Fabricius), Xylotrechus aceris Fisher, Xylotrechus sagittatus sagittatus (Germar), Tylonotus bimaculatus Haldeman, Lepturges angulatus (LeConte), Lepturges symmetricus (Haldeman), Urgleptes querci (Fitch), Oplosia nubila (LeConte), Eupogonius subarmatus (LeConte), Monochamus carolinensis (Olivier), and Pogonocherus parvulus LeConte. Urgleptes signatus (LeConte) and Urgleptes querci are newly recorded from Nova Scotia. All but two specimens were collected in 12-funnel Lindgren traps. Xylotrechus aceris, Tylonotus bimaculatus, Lepturges angulatus, Lepturges symmetricus, Urgleptes signatus (NS), and Pogonocherus parvulus were detected exclusively in traps deployed in the forest canopy, and most individuals of Oplosia nubila and Monochamus carolinensis were captured in canopy traps. Arhopalus obsoletus, Atimia confusa confusa, Callidium frigidum, Phymatodes testaceus, and Xylotrechus sagittatus sagittatus were captured almost exclusively in traps near (1 m above) the forest floor. These results highlight the importance of sampling both the understory and upper canopy when using traps for surveying diversity of Cerambycidae.