New data on the distribution of the ground beetle <Emphasis Type="Italic">Eremosphodrus</Emphasis> (s. str.) <Emphasis Type="Italic">rotundicollis</Emphasis> (Reitter, 1894) (Coleoptera,Carabidae)

Data on the distribution of Eremosphodrus (s. str.) rotundicollis (Reitter, 1894) are discussed. The species is recorded for the territory of Uzbekistan for the first time (Termez District of Surkhan-Darya Province).     

Species of the subgenus <Emphasis Type="Italic">Baudia</Emphasis> of the genus <Emphasis Type="Italic">Badister</Emphasis> (Coleoptera,Carabidae) from the southern Sikhote-Alin Mountains

Data on two species of the subgenus Baudia Ragusa, 1884, B. ishigakiensis Habu, 1975 and B. marginellus Bates, 1873 new to the fauna of the southern Sikhote-Alin Mountains, are presented. A key to species of this subgenus is provided. Holotypes of B. nigriceps A. Morawitz, 1863 and B. ussuriensis Jedli?ka, 1937 are designated. New data on the distribution of the species in Eastern Asia are given.     

New combinations and typifications in <Emphasis Type="Italic">Bistorta</Emphasis>, <Emphasis Type="Italic">Persicaria</Emphasis>, <Emphasis Type="Italic">Polygonum,</Emphasis> and <Emphasis Type="Italic">Rumex</Emphasis> (Polygonaceae)

New combinations are proposed in anticipation of the Polygonaceae treatment in the forthcoming volume of Intermountain Flora: Polygonum kelloggii var. esotericum, P. kelloggii var. watsonii , Rumex densiflorus var. pycnanthus , R. salicifolius var. utahensis, and R. occidentalis var. tomentellus. Typifications are proposed to facilitate ongoing studies in Polygonaceae and to maintain current usage.     

A review of the subgenus <Emphasis Type="Italic">Carpelimus</Emphasis> s. str. (Coleoptera,Staphylinidae) from tropical Africa

A review of the subgenus Carpelimus (s. str.) from tropical Africa is given. The subgenus includes 10 species. A new species C. uhligi sp. n. is described, neotype of Trogophloeus insularis, Kraatz 1858 is designated. T. aequithorax Bernhauer, 1932 is placed in synonymy with C. dieganus (Fauvel, 1904); T. oculatus Wollaston, 1865 and T. meridioafricanus Scheerpeltz, 1974, with C. insularis (Kraatz, 1858), T. rudebecki Scheerpeltz, 1974 with C. memnonius (Erichson, 1840); T. yemenicus Coiffait, 1981 with C. niloticus (Erichson, 1840); T. nigerrimus Coiffait, 1935 and T. mimus Cameron, 1945, with C. rufitarsis (Fauvel, 1907); and T. bredoi Bernhauer, 1943, with C. transmarinus (Fauvel, 1907). Lectotypes of T. aequithorax, T. calidus, T. nigerrimus, and T. bredoi are designated. Twenty species are transferred from the genus Carpelimus, and the following new combinations are formed: Thinodromus brincki Scheerpeltz, 1972; Th. montiumdraconis Scheerpeltz, 1974; Th. rhodesianus Scheerpeltz, 1974; and Th. sudanensis Scheerpeltz, 1974 comb. nn.). The initial generic placement of Carpelimus luzidus Cameron, 1944 is restored.     

Isolation and <Emphasis Type="Italic">S</Emphasis>-genotyping application of <Emphasis Type="Italic">S</Emphasis>-allelic polymorphic <Emphasis Type="Italic">MdSLFB</Emphasis>s in apple (<Emphasis Type="Italic">Malus domestica</Emphasis> Borkh.)

Apple (Malus domestica Borkh.) possesses gametophytic self-incompatibility (GSI) which is controlled by S-RNase in the pistil as well as a pollen S-determinant that has not been well characterized. The identification of S-locus F-box brother (SFBB) genes, which are good candidates for the pollen S-determinant in apple and pear, indicated the presence of multiple S-allelic polymorphic F-box genes at the S-locus. In apple, two SFBB gene groups have been described, while there are at least three groups in pear. In this report, we identified five MdSLFB (S-RNase-linked F-box) genes from four different S-genotypes of apple. These genes showed pollen- and S-allele-specific expression with a high polymorphism among S-alleles. The phylogenetic tree suggested that some of them belong to SFBBα or β groups as described previously, while others appear to be different from SFBBs. In particular, the presence of MdSLFB3 and MdSLFB9 suggested that there are more S-allelic polymorphic F-box gene groups in the S-locus besides α and β. Based on the sequence polymorphism of MdSLFBs, we developed an S-genotyping system for apple cultivars. In addition, we isolated twelve MdSLFB-like genes, which showed pollen-specific expression without S-allelic polymorphism.     

<Emphasis Type="Italic">Viola woosanensis</Emphasis>, a recurrent spontaneous hybrid between <Emphasis Type="Italic">V. ulleungdoensis</Emphasis> and <Emphasis Type="Italic">V. chaerophylloides</Emphasis> (Violaceae) endemic to Ulleung Island,Korea

Ulleung Island is an oceanic volcanic island in Korea, which has never been connected to the adjacent continent. Previous studies highlighted Ulleung Island as an excellent system to study the pattern and process of early stages of flowering plant evolutions on oceanic island. The predominant mode of speciation in flowering plants on Ulleung Island appears to be anagenesis. However, the potentially important role of hybrid speciation among incompletely reproductively isolated lineages cannot be ruled out. Viola woosanensis (Violaceae) is of purportedly hybrid origin between V. ulleungdoensis (i.e., formerly recognized as V. selkirkii in Ulleung Island) and V. chaerophylloides, based on morphology. To examine the origin of V. woosanensis, we sampled a total of 80 accessions, including V. woosanensis and its putative parental species and sequenced nrDNA ITS, and four highly variable chloroplast noncoding regions (trnL-trnF, rpl16 intron, atpF-atpH, and psbA-trnH). Representative species of Viola from Korea were also included in the phylogenetic analyses (maximum parsimony, maximum likelihood, and Bayesian inference). Additive polymorphic sites in the nrDNA ITS regions were confirmed by cloning amplicons from representative species. The molecular data strongly supported the hybrid origin of V. woosanensis, and the maternal and paternal parent were determined to be V. ulleungdoensis and V. chaerophylloides, respectively. The presence of two parental ribotypes in V. woosanensis (with the exception in one population) was confirmed by cloning, suggesting V. woosanensis is primarily the F₁ generation. No trace of backcrossing and introgression with its parents was detected due to low fertility of hybrid species. We found a multiple and unidirectional hybrid origin of V. woosanensis. Additional studies are required to determine which factors contribute to asymmetric gene flow of Viola species in Ulleung Island.     

Taxonomy and geographic distribution of <Emphasis Type="Italic">Carex lucorum</Emphasis> var. <Emphasis Type="Italic">austrolucorum</Emphasis> (section <Emphasis Type="Italic">Acrocystis</Emphasis>, Cyperaceae)

Carex lucorum currently comprises two morphologically similar varieties. We revise the taxonomy of C. lucorum, and map the distribution of the taxa. Based on multivariate analyses and analyses of variance of measured characters, allopatry, and previous studies revealing differences in achene micromorphology, chromosome differences, and flavonoid chemistry, we elevate C. lucorum var. austrolucorum to species status as C. austrolucorum. We provide an identification key to C. lucorum and its closest relatives.     

<Emphasis Type="Italic">Ty</Emphasis>1<Emphasis Type="Italic">/copia</Emphasis>- and <Emphasis Type="Italic">Ty</Emphasis>3<Emphasis Type="Italic">/gypsy</Emphasis>-like DNA sequences in <Emphasis Type="Italic">Helianthus </Emphasis>species

Two repeated DNA sequences isolated from a partial genomic DNA library of Helianthus annuus, p HaS13 and p HaS211, were shown to represent portions of the int gene of a Ty3 /gypsy retroelement and of the RNase-Hgene of a Ty1 /copia retroelement, respectively. Southern blotting patterns obtained by hybridizing the two probes to BglII- or DraI-digested genomic DNA from different Helianthus species showed p HaS13 and p HaS211 were parts of dispersed repeats at least 8 and 7 kb in length, respectively, that were conserved in all species studied. Comparable hybridization patterns were obtained in all species with p HaS13. By contrast, the patterns obtained by hybridizing p HaS211 clearly differentiated annual species from perennials. The frequencies of p HaS13- and p HaS211-related sequences in different species were 4.3x10(4)-1.3x10(5) copies and 9.9x10(2)-8.1x10(3) copies per picogram of DNA, respectively. The frequency of p HaS13-related sequences varied widely within annual species, while no significant difference was observed among perennial species. Conversely, the frequency variation of p HaS211-related sequences was as large within annual species as within perennials. Sequences of both families were found to be dispersed along the length of all chromosomes in all species studied. However, Ty3 /gypsy-like sequences were localized preferentially at the centromeric regions, whereas Ty1/ copia-like sequences were less represented or absent around the centromeres and plentiful at the chromosome ends. These findings suggest that the two sequence families played a role in Helianthusgenome evolution and species divergence, evolved independently in the same genomic backgrounds and in annual or perennial species, and acquired different possible functions in the host genomes.     

<Emphasis Type="Italic">Agrobacterium tumefaciens</Emphasis>-mediated transformation of <Emphasis Type="Italic">Festuca arundinacea</Emphasis> (Schreb.) and <Emphasis Type="Italic">Lolium multiflorum</Emphasis> (Lam.)

Agrobacterium tumefaciens strain LBA4404 carrying plasmid pTOK233 encoding the hygromycin resistance (hph) and beta-glucuronidase (uidA) genes has been used to transform two agronomic grass species: tall fescue (Festuca arundinacea) and Italian ryegrass (Lolium multiflorum). Embryogenic cell suspension colonies or young embryogenic calli were co-cultured with Agrobacterium in the presence of acetosyringone. Colonies were grown under hygromycin selection with cefotaxime and surviving colonies plated on embryogenesis media. Eight Lolium (six independent lines) and two Festuca plants (independent lines) were regenerated and established in soil. All plants were hygromycin-resistant, but histochemical determination of GUS activity showed that only one Festuca plant and one Lolium plant expressed GUS. Three GUS-negative transgenic L. multiflorum and the two F. arundinacea plants were vernalised and allowed to flower. All three Lolium plants were male- and female-fertile, but the Festuca plants failed to produce seed. Progeny analysis of L. multiflorum showed a 24-68% inheritance of the hph and uidA genes in the three lines with no significant difference between paternal and maternal gene transmission. However, significant differences were noted between the paternal and maternal expression of hygromycin resistance.     

A new species of <Emphasis Type="Italic">Macroptilium</Emphasis> (Benth.) Urb. (<Emphasis Type="Italic">Leguminosae</Emphasis>: <Emphasis Type="Italic">Papilionoideae</Emphasis>: <Emphasis Type="Italic">Phaseolinae</Emphasis>) from North-Eastern Brazil

Summary  A new species of Macroptilium sect. Microcochle (Benth.) J. A. Lackey is herein described from the states of Piauí and Bahia, Brazil. Macroptilium cochleatum is characterized by few-flowered inflorescences, calyx teeth longer than the tube, a tightly four-coiled keel, and linear, patent fruits. The discovery of this unique Macroptilium species, with its distally coiled keel, expands the diagnostic features of the genus. A key to the Brazilian species of sect. Microcochle is provided.     

Factors affecting <Emphasis Type="Italic">Agrobacterium</Emphasis>-mediated transformation in <Emphasis Type="Italic">Hybanthus</Emphasis><Emphasis Type="Italic">enneaspermus</Emphasis> (L.) F. Muell.

Agrobacterium tumefaciens-mediated transformation system was established for Hybanthus enneaspermus using leaf explants with the strain LBA4404 harbouring pCAMBIA 2301 carrying the nptII and gusA genes. Sensitivity of leaf explants to kanamycin was standardized (100 mg/l) for screening the transgenic plants. Transformation parameters (OD, virulence inducer, infection time, co-cultivation period, bactericidal antibiotics, etc.) influencing the gene transfer and integration were assessed in the present investigation. Fourteen-day pre-cultured explants were subjected with Agrobacterium strain LBA4404. Optimized parameters such as culture density of 0.5 OD₆₀₀, infection time of 6 min, AS concentration of 150 µM with 3 days co-cultivation revealed maximum transformation efficiency based on GUS expression assay. The presence of gusA in transgenics was confirmed by polymerase chain reaction and Southern blotting analysis. The present transformation experiment yielded 20 shoots/explant with higher transformation efficiency (28 %). The protocol could be used to introduce genes for trait improvement as well as for altering metabolic pathway for secondary metabolites production.     

Accompanying vegetation in young <Emphasis Type="Italic">Pinus radiata</Emphasis> plantations enhances recolonization by <Emphasis Type="Italic">Ceroglossus chilensis</Emphasis> (Coleoptera: Carabidae) after clearcutting

The replacement of native forests by Pinus radiata plantations modifies habitat availability and quality for wildlife, constituting a threat to species survival. However, the presence of understory in mature pine plantations minimizes the negative impacts of native forest replacement, rendering a secondary habitat for wildlife. Whether forest-dwelling species recolonize clear-felled areas pending on the spontaneous development of accompanying vegetation growing after harvesting is yet to be assessed. In this context, we analyze the abundance, movement and habitat selection of the endemic ground beetle Ceroglossus chilensis (Coleoptera: Carabidae) in an anthropic forest landscape consisting of native forest remnants, adult pine plantations (>?20 years) with a well-developed understory, and young (1–2 years) pine plantations with varying degrees of accompanying vegetation development. Particularly, we analyze the likelihood that C. chilensis would recolonize young pine plantations depending on the presence (>?70% cover) or the absence (<?20% cover) of this accompanying vegetation. C. chilensis shows a greater probability of selecting habitats with understory (pine plantations and native forest) and young plantations with accompanying vegetation (future understory) than habitats without such vegetation. Movement of C. chilensis also favors their permanence in habitats with understory vegetation, coinciding with higher abundances than in young pine plantations devoid of accompanying vegetation. Hence, the effect of clearcutting could be mitigated by allowing the development of accompanying vegetation into a future understory, which facilitates the recolonization of pine plantations and its use as secondary habitat for wildlife.     

Phylogenetic Relationships within <Emphasis Type="Italic">Phyllophaga</Emphasis> Harris (sensu <Emphasis Type="Italic">lato</Emphasis>) (Coleoptera: Melolonthidae,Melolonthinae) with Emphasis on <Emphasis Type="Italic">Listrochelus</Emphasis> Blanchard

As partial results of a long-term project for the revision of the supraspecific classification of the American Melolonthini, using phylogenetic methods, interesting information on the relationships between the subgeneric groups of Phyllophaga Harris proposed by Saylor were obtained. The genus Listrochelus was described by Blanchard in 1851. However, in 1940, Saylor reduced it as a subgenus of Phyllophaga. The objective of the present study is to confirm the monophyly of Listrochelus by means of a phylogenetic analysis. A total of 132 species were analyzed; 31 species of them belong to Listrochelus, 76 are from other groups of Phyllophaga, and 25 species are from the outgroup. A morphological matrix with 281 characters was codified. A traditional search with 1000 iterations was performed in TNT. Branch support was investigated using the bootstrap method. Parsimony analysis resulted in ten equally parsimonious trees. The topology obtained in the strict consensus tree shows that the limits of Listrochelus are very clear (bootstrap 99%), supported by five synapomorphies and a combination of eight character states that are not exclusive to the group. Based on the hypothesis obtained, the restitution of the genus Listrochelus is proposed.     

<Emphasis Type="Italic">In silico</Emphasis> and <Emphasis Type="Italic">in situ</Emphasis> characterization of the zebrafish (<Emphasis Type="Italic">Danio rerio</Emphasis>) <Emphasis Type="Italic">gnrh3</Emphasis> (sGnRH) gene

Background

Gonadotropin releasing hormone (GnRH) is responsible for stimulation of gonadotropic hormone (GtH) in the hypothalamus-pituitary-gonadal axis (HPG). The regulatory mechanisms responsible for brain specificity make the promoter attractive for in silico analysis and reporter gene studies in zebrafish (Danio rerio).

Results

We have characterized a zebrafish [Trp⁷, Leu⁸] or salmon (s) GnRH variant, gnrh 3. The gene includes a 1.6 Kb upstream regulatory region and displays the conserved structure of 4 exons and 3 introns, as seen in other species. An in silico defined enhancer at -976 in the zebrafish promoter, containing adjacent binding sites for Oct-1, CREB and Sp1, was predicted in 2 mammalian and 5 teleost GnRH promoters. Reporter gene studies confirmed the importance of this enhancer for cell specific expression in zebrafish. Interestingly the promoter of human GnRH-I, known as mammalian GnRH (mGnRH), was shown capable of driving cell specific reporter gene expression in transgenic zebrafish.

Conclusions

The characterized zebrafish Gnrh3 decapeptide exhibits complete homology to the Atlantic salmon (Salmo salar) GnRH-III variant. In silico analysis of mammalian and teleost GnRH promoters revealed a conserved enhancer possessing binding sites for Oct-1, CREB and Sp1. Transgenic and transient reporter gene expression in zebrafish larvae, confirmed the importance of the in silico defined zebrafish enhancer at -976. The capability of the human GnRH-I promoter of directing cell specific reporter gene expression in zebrafish supports orthology between GnRH-I and GnRH-III.

     

Molecular Phylogenetics and Biogeography of the Caribbean-Centered <Emphasis Type="Italic">Croton</Emphasis> Subgenus <Emphasis Type="Italic">Moacroton</Emphasis> (Euphorbiaceae s.s.)

Initial molecular phylogenetic studies established the monophylly of the large genus Croton (Euphorbiaceae s.s.) and suggested that the group originated in the New World. A denser and more targeted sampling of Croton species points to a South American origin for the genus. The nuclear and chloroplast genomes indicate a different rooting for the phylogeny of Croton. Although we favor the rooting indicated by the chloroplast data our conclusions are also consistent with the topology inferred from the nuclear data. The satellite genera Cubacroton and Moacroton are embedded within Croton. These two genera are synonimized into Croton and a new subgenus, Croton subgenus Moacroton, is circumscribed to include them and their allied Croton species. Croton subgenus Moacroton is morphologically characterized by a primarily lepidote indumentum, bifid or simple styles, and pistillate flowers with sepals that are connate at the base. This newly circumscribed subgenus is found from North America to South America, and in contrast to the majority of Croton species most of its members are found in mesic habitats. The group is most diverse in the greater Caribbean basin. A molecular clock was calibrated to the phylogeny using the available Euphorbiaceae fossils. The timing and pattern of diversification of Croton is consistent with both the GAARlandia and Laurasian migration hypotheses. A single species, Croton poecilanthus from Puerto Rico, is placed incongruently by its nuclear and chloroplast genomes. The possibility of this species being of hybrid origin is discussed.      

<Emphasis Type="Italic">Entamoeba histolytica</Emphasis> and <Emphasis Type="Italic">E. dispar</Emphasis> infections in captive macaques (<Emphasis Type="Italic">Macaca fascicularis</Emphasis>) in the Philippines

Entamoeba histolytica is a protozoan parasite that infects man and animals. This parasite has a global distribution and the disease it causes is usually characterized by diarrhea. In order to detect the parasite, it is necessary to differentiate it from Entamoeba dispar. E. dispar appears morphologically similar to E. histolytica but does not cause disease and tissue invasion. This study reports on the prevalence of E. histolytica and E. dispar among captive macaques in a primate facility in the Philippines. PCR was used to correctly identify both Entamoeba species. Indirect fluorescent antibody test (IFAT) was also performed to determine the seroprevalence of amebiasis in the captive macaques. Based on PCR targeting of the peroxiredoxin gene, of the 96 stool samples collected, 23 (24%) contained E. histolytica while 32 (33%) contained E. dispar. IFAT revealed 26 (27%) serum samples positive for antibodies against E. histolytica. Sequence analysis of the 18S rRNA gene showed that the 23 E. histolytica isolates were identical to human E. histolytica isolates deposited in the GenBank and not Entamoeba nuttalli as found in macaques in other recent reports. The Philippines is a major exporter of monkeys for biomedical research purposes, so screening animals before transporting them to other locations lessens the risk of spreading zoonoses to a wider area. This is the first report of the molecular detection of E. histolytica and E. dispar among macaques in the Philippines. This study complements the limited information available on the animal hosts of E. histolytica in the Philippines.     

Pedogamy in <Emphasis Type="Italic">Neidium</Emphasis> (<Emphasis Type="Italic">Bacillariophyceae</Emphasis>)

Single (unpaired) vegetative cells of freshwater pennate diatom Neidium cf. ampliatum differentiated into gametangia and produced a single zygote (auxospore) via a pedogamic process. The gametic nuclei fused after auxospore expansion had begun. The auxospore expanded in parallel to the apical axis of the gametangium.     

Floral development and systematic position of <Emphasis Type="Italic">Arillastrum</Emphasis>, <Emphasis Type="Italic">Allosyncarpia</Emphasis>, <Emphasis Type="Italic">Stockwellia</Emphasis> and <Emphasis Type="Italic">Eucalyptopsis</Emphasis> (Myrtaceae)

We have investigated the floral ontogeny of Arillastrum, Allosyncarpia, Stockwellia and Eucalyptopsis (of the eucalypt group, Myrtaceae) using scanning electron microscopy and light microscopy. Several critical characters for establishing relationships between these genera and to the eucalypts have been determined. The absence of compound petaline primordia in Arillastrum, Allosyncarpia, Stockwellia and Eucalyptopsis excludes these taxa from the eucalypt clade. Post-anthesis circumscissile abscission of the hypanthium above the ovary in Stockwellia, Eucalyptopsis and Allosyncarpia is evidence that these three taxa form a monophyletic group; undifferentiated perianth parts and elongated fusiform buds are characters that unite Stockwellia and Eucalyptopsis as sister taxa. No floral characters clearly associate Arillastrum with either the eucalypt clade or the clade of Stockwellia, Eucalyptopsis and Allosyncarpia.We gratefully acknowledge Clyde Dunlop and Bob Harwood (Northern Territory Herbarium) for collecting specimens of Allosyncarpia, and Bruce Gray (Atherton) for collecting specimens of Stockwellia. The Australian National Herbarium (CANB) kindly lent herbarium specimens of Eucalyptopsis for examination. This research was supported by a University of Melbourne Research Development Grant to Andrew Drinnan.     

<Emphasis Type="Italic">Agrobacterium</Emphasis><Emphasis Type="Italic">tumefaciens</Emphasis>-mediated transformation of callus cells of <Emphasis Type="Italic">Crataegus</Emphasis><Emphasis Type="Italic">aronia</Emphasis>

A genetic transformation system has been developed for callus cells of Crataegus aronia using Agrobacterium tumefaciens. Callus culture was established from internodal stem segments incubated on Murashige and Skoog (MS) medium supplemented with 5 mg l⁻¹ Indole-3-butyric acid (IBA) and 0.5 mg l⁻¹ 6-benzyladenine (BA). In order to optimize the callus culture system with respect to callus growth and coloration, different types and concentrations of plant growth regulators were tested. Results indicated that the best average fresh weight of red colored callus was obtained on MS medium supplemented with 2 mg l⁻¹ 2,4-dichlorophenoxyacetic acid (2,4-D) and 1.5 mg l⁻¹ kinetin (Kin) (callus maintenance medium). Callus cells were co-cultivated with Agrobacterium harboring the binary plasmid pCAMBIA1302 carrying the mgfp5 and hygromycin phosphotransferase (hptII) genes conferring green fluorescent protein (GFP) activity and hygromycin resistance, respectively. Putative transgenic calli were obtained 4 weeks after incubation of the co-cultivated explants onto maintenance medium supplemented with 50 mg l⁻¹ hygromycin. Molecular analysis confirmed the integration of the transgenes in transformed callus. To our knowledge, this is the first time to report an Agrobacterium-mediated transformation system in Crataegus aronia.