Development and characterization of 12 compound microsatellite markers in <Emphasis Type=Italic>Platypus </Emphasis><Emphasis Type=Italic>quercivorus</Emphasis> (Murayama) (Coleoptera: Platypodidae)

Twelve compound microsatellite loci were isolated from the ambrosia beetle Platypus quercivorus. Every locus was polymorphic among 50 individuals from two localities, with two to six alleles per locus, without linkage disequilibrium. The observed and expected heterozygosities ranged from 0.140 to 0.740 and from 0.255 to 0.731, respectively. These markers will be available for population genetic studies and parentage analysis of this beetle.     

Isolation and characterization of microsatellite loci in the Brazilian orchid <Emphasis Type=Italic>Epidendrum fulgens</Emphasis>

Epidendrum fulgens has a patchy distribution along the Atlantic Rainforest in the Brazilian coast, due to the destruction of its native habitat. Here, we report on both the development of nine new microsatellite markers isolated from this species and the characterization of their allele variability in two distant and unrelated populations. The number of alleles observed for each locus ranged from 2 to 17 with an average of 6.4 alleles per locus. These microsatellites should be valuable tools for studying the effect of habitat fragmentation on the genetic structure of E. fulgens populations.     

Thidiazuron Promotes <Emphasis Type=Italic>In Vitro</Emphasis> Plant Regeneration of <Emphasis Type=Italic>Arachis correntina</Emphasis> (<Emphasis Type=Italic>Leguminosae</Emphasis>) via Organogenesis

Arachis correntina (Burkart) Krapov. & W.C. Gregory is a herbaceous perennial leguminous plant growing in the Northeast of the Province Corrientes, Argentina. It is important as forage. The development of new A. correntina cultivars with improved traits could be facilitated through the application of biotechnological strategies. The purpose of this study was to investigate the plant regeneration potential of mature leaves of A. correntina in tissue culture. Buds were induced from both petiole and laminae on 0.7% agar-solidified medium containing 3% sucrose, salts and vitamins from Murashige and Skoog (MS) supplemented with 0.5–25 M thidiazuron (TDZ). Shoot induction was achieved by transference of calli with buds to MS supplemented with 5 M TDZ. Fifty-four percent of the regenerated shoot rooted on MS + 5 M naphthaleneacetic acid. Histological studies revealed that shoots regenerated via organogenesis.     

Development of tetranucleotide microsatellite loci for the finless porpoise (<Emphasis Type=Italic>Neophocaena phocaenoides</Emphasis>)

The finless porpoise, Neophocaena phocaenoides, is endemic to the coastal waters of the Indo-Pacific, ranging from the Persian Gulf to Japan. Nine tetranucleotide microsatellite loci were isolated from the finless porpoise (Neophocaena phocaenoides). Polymorphism of each locus was assessed in 39 unrelated individuals from the Yellow Sea and the South China Sea of the Chinese waters. The number of alleles per locus varied from 2 to 11. The ranges of observed and expected heterozygosity were 0.154–0.795 and 0.146–0.839, respectively. Cross-species amplification of these loci was tested in other cetacean species. These microsatellite markers described here will be suitable for population genetic studies of finless porpoises and other cetacean species.     

Twelve polymorphic microsatellite loci for <Emphasis Type="Italic">Achnatherum inebrians</Emphasis> (Poaceae)

Microsatellites are often highly variable and abundant in most complex genomes, therefore are widely used in population genetic studies. In this study, twelve polymorphic microsatellite loci were isolated and characterized for the Achnatherum inebrians, a plant abundant in grasslands of Northwest China. Characterization of 24 A. inebrians individuals form four geographically distant populations (Gansu, Qinghai, Xinjiang and Inner Mongolia provinces) showed moderate to high allelic diversity ranging from 3 to 13 alleles per locus, and the expected heterozygosity ranging from 0.41 to 0.67. No evidence of linkage disequilibrium was found for any locus pairwise comparisons. The markers described here will be useful for investigating the genetic diversity, genetic structure and gene flow of this species. Na Chen and Yan-Zhuo Yang contributed equally to this work.     

A new name in <Emphasis Type=Italic>Pavetta</Emphasis> L. (<Emphasis Type=Italic>Rubiaceae</Emphasis>)

Summary  The name Pavetta modesta (Hiern) S. E. Dawson is a later homonym of P. modesta Bremek. Pavetta crystalensis is proposed as a new name.     

<Emphasis Type=Italic>In vitro</Emphasis> propagation of ten threatened species of <Emphasis Type=Italic>Mammillaria</Emphasis> (Cactaceae)

Mammillaria species are the most numerous within Cactaceae family, and some of them are threatened with extinction as a result of human activities. In this work, results of in vitro propagation are presented for ten Mammillaria species, testing 20 combinations of indole-3-acetic acid (IAA) and kinetin. Best results on shoot formation were obtained using kinetin at two levels: 27.9 and 46.5 μM. All IAA levels tested were able to induce de novo shoot formation in M. bocasana, M. densispina, M. hahniana, M. hutchisoniana, M. orcutii, M. pectinifera, M. perbella, M. picta, M. rhodantha, and M. zephyranthoides. Depending on the IAA level tested, four responding groups were observed concerning their highest shoot-formation number. For all species, the highest average of shoot formation was achieved with 5.7:46.5 or 11.4:46.5 μM IAA/kinetin, yielding 4.8 and 4.7 shoots per explant, respectively, in 60 d. Rooting of regenerated shoots was achieved by leaving the explants in their shoot-induction medium or transferring them to half-strength MS medium. Hardening of regenerated plants was successfully achieved by planting them in peat moss substrate after a desiccation treatment at room temperature for 3 d.     

<Emphasis Type=Italic>Cardamine plumieri</Emphasis> (<Emphasis Type=Italic>Brassicaceae</Emphasis>) confirmed to the Flora of Serbia

The south-European Cardamine plumieri Vill. (sect. Pteroneurum subsect. Cryptopterum) is confirmed for the Flora of Serbia. Several populations were discovered in the central and western part of country, exclusively on the serpentine bedrocks. Morphological, chorological and ecological characteristics of C. plumieri was studied.     

Development of 37 microsatellite loci for the great gray owl (<Emphasis Type=Italic>Strix nebulosa</Emphasis>) and other <Emphasis Type=Italic>Strix</Emphasis> spp. owls

We developed 37 great gray owl (Strix nebulosa) microsatellite primers from CA and TAGA enriched genomic libraries. Primers were tested in 15 great gray owls from California, USA and Alberta, Canada as well as two other Strix species, spotted owl (S. occidentalis) and barred owl (S. varia). These markers will have broad application in investigations of Strix population structure and genetic diversity.     

Typification of <Emphasis Type=Italic>Metrosideros regelii</Emphasis> (<Emphasis Type=Italic>Myrtaceae</Emphasis>) and consideration of its generic position

Summary  The type specimen of Metrosideros regelii is discussed. It contains a mixture of two species, representing different genera, and a lectotype is chosen. The generic position of the species is considered in the light of morphology and recent molecular evidence and the new combination, Mearnsia regelii, made.     

Microbiological and physicochemical factors affecting <Emphasis Type=Italic>Aspergillus</Emphasis> section <Emphasis Type=Italic>Flavi</Emphasis> incidence in Cavendish banana (<Emphasis Type=Italic>Musa cavendishii</Emphasis>) chips production in Southern Philippines

Microbiological and physicochemical factors affecting the incidence of Aspergillus section Flavi in dried Cavendish banana (Musa cavendishii) chips production in Southern Philippines were examined. The average counts of Aspergillus section Flavi (AFC) in fresh and dried Cavendish bananas from 10 production batches of the Philippine Agro-Industrial Development Cooperative in Davao del Norte, Southern Philippines were 1.2 × 10² and 1.6 × 10² cfu/g, respectively. Isolates from both samples were identified to be Aspergillus flavus based on spore type and conidial structure of isolates. An increasing trend in the AFC of Cavendish bananas was observed during dried banana chips processing. Variability in the AFC between production batches was attributed to differences in aerobic and fungal populations and physicochemical characteristics of the fruits, peel damage of the raw materials, concentration of AFC in the air and food-contact surfaces of the production area, and temperature and relative humidity (RH) conditions of the environment during production and storage. Physicochemical characteristics of Cavendish bananas from the receipt of raw materials up to the first day of drying were within the reported range of values allowing growth and toxin production by aflatoxigenic fungi. Air-borne AFC varied depending on the section of the production area examined. The close proximity of the waste disposal area from the production operation to the preparation, drying and storage areas suggests that cross-contamination, probably air-borne or insect-borne was a likely occurrence. The hands of workers were also identified as AFC sources. Results of this study highlight the need for the development of strategies to control aflatoxigenic fungi and aflatoxin contamination in Philippine dried Cavendish bananas.     

The expression of analgesic-antitumor peptide (<Emphasis Type=Italic>AGAP</Emphasis>) from Chinese <Emphasis Type=Italic>Buthus</Emphasis><Emphasis Type=Italic>martensii</Emphasis> Karsch in transgenic tobacco and tomato

The present study aimed to obtain analgesic-antitumor peptide (AGAP) gene expression in plants. The analgesic-antitumor peptide (AGAP) gene was from the venom of Buthus martensii Karsch. Previous studies showed that AGAP has both analgesic and antitumor activities, suggesting that AGAP would be useful in clinical situations as an antitumor drug. Given that using a plant as an expression vector has more advantages than prokaryotic expression, we tried to obtain transgenic plants containing AGAP. In the present study, the AGAP gene was cloned into the plasmid pBI121 to obtain the plant expression vector pBI-AGAP. By tri-parental mating and freeze–thaw transformation, pBI-AGAP was transformed into Agrobacterium tumefaciens LBA4404. Tobacco (Nicotiana tabacum) and tomato (Lycopersicom esculentum) were transformed by the method of Agrobacterium-mediated leaf disc transformation. The transformants were then screened to grow and root on media containing kanamycin. Finally, transformations were confirmed by analysis of PCR, RT-PCR and western blotting. The results showed that the AGAP gene was integrated into the genomic DNA of tobacco and tomato and was successfully expressed. Therefore, the present study suggests a potential industrial application of AGAP expressed in plants.     

Social encapsulation of the small hive beetle (<Emphasis Type=Italic>Aethina tumida</Emphasis> Murray) by European honeybees (<Emphasis Type=Italic>Apis mellifera</Emphasis> L.)

Summary European and African subspecies of honeybees (Apis mellifera L.) utilize social encapsulation to contain the small hive beetle (Aethina tumida Murray), a honeybee colony scavenger. Using social encapsulation, African honeybees successfully limit beetle reproduction that can devastate host colonies. In sharp contrast, European honeybees often fail to contain beetles, possibly because their social encapsulation skills may be less developed than those of African honeybees. In this study, we quantify beetle and European honeybee behaviours associated with social encapsulation, describe colony and time (morning and evening) differences in these behaviours (to identify possible circadian rhythms), and detail intra-colonial, encapsulated beetle distributions. The data help explain the susceptibility of European honeybees to depredation by small hive beetles. There were significant colony differences in a number of social encapsulation behaviours (the number of beetle prisons and beetles per prison, and the proportion of prison guard bees biting at encapsulated beetles) suggesting that successful encapsulation of beetles by European bees varies between colonies. We also found evidence for the existence of circadian rhythms in small hive beetles, as they were more active in the evening rather than morning. In response to increased beetle activity during the evening, there was an increase in the number of prison guard bees during evening. Additionally, the bees successfully kept most (~93%) beetles out of the combs at all times, suggesting that social encapsulation by European honeybees is sufficient to control small populations of beetles (as seen in this study) but may ultimately fail if beetle populations are high.Received 20 January 2003; revised 21 April 2003; accepted 29 April 2003.     

Micromorphological differences between some European and American <Emphasis Type=Italic>Fraxinus</Emphasis> (<Emphasis Type=Italic>Oleaceae</Emphasis>) species

Micromorphological differences in leaves and pollen between two American (Fraxinus americana L., F. pennsylvanica Marshall) and two European (F. angustifolia Vahl, F. excelsior L.) ash species were studied using scanning electron microscope. The types, dimensions and distribution of characteristic trichomes were established and measured. Capitate hairs on the leaves had the same shape in all researched ash species. Acicular hairs were regularly present in two American ash species, but very rarely in the glabrous phase of F. angustifolia and F. excelsior. Only F. americana had coronulate abaxial surface of leaves. Pollen of F. angustifolia and F. excelsior had 3 (tricolpate) apertures, and F. americana and F. pennsylvanica 4 (stephanocolpate) apertures. Based on the appearance of the reticulum it’s possible to clearly distinguish all four species. F. angustifolia and F. pennsylvanica had muri with transversal ridges and seldom granules. Muri of F. excelsior and F. americana had slightly visible transversal ridges, and because of that noticeable granules.     

Genetic Distance in Housekeeping Genes Between <Emphasis Type=Italic>Plasmodium falciparum</Emphasis> and <Emphasis Type=Italic>Plasmodium reichenowi</Emphasis> and Within <Emphasis Type=Italic>P. falciparum</Emphasis>

The time to the most recent common ancestor of the extant populations of Plasmodium falciparum is controversial. The controversy primarily stems from the limited availability of sequences from Plasmodium reichenowi, a chimpanzee malaria parasite closely related to P. falciparum. Since the rate of nucleotide substitution differs in different loci and DNA regions, the estimation of genetic distance between P. falciparum and P. reichenowi should be performed using orthologous sequences that are evolving neutrally. Here, we obtained full-length sequences of two housekeeping genes, sarcoplasmic and endoplasmic reticulum Ca²⁺-ATPase (serca) and lactate dehydrogenase (ldh), from 11 isolates of P. falciparum and 1 isolate of P. reichenowi and estimate the interspecific genetic distance (divergence) between the two species and intraspecific genetic distance (polymorphism) within P. falciparum. Interspecific distance and intraspecific distance at synonymous sites of interspecies-conserved regions of serca and ldh were 0.0672±0.0088 and 0.0011±0.0007, respectively, using the Nei and Gojobori method. Based on the ratio of interspecific distance to intraspecific distance, the time to the most recent common ancestor of P. falciparum was estimated to be (8.30±5.40) × 10⁴ and (11.62±7.56) × 10⁴ years ago, assuming the divergence time of the two parasite species to be 5 and 7 million years ago, respectively.This article contains an online supplementary table.Reviewing Editor: Dr. Martin Kreitman     

Karyotype analysis of medicinal plant <Emphasis Type=Italic>Liriope spicata</Emphasis> var. <Emphasis Type=Italic>prolifera</Emphasis> (Liliaceae)

Karyotype of Liriope spicata var. prolifera, a Chinese endemic species, was described in detail for the first time. Its proto-variety L. spicata was also investigated for comparison. The basic chromosome number of these two species was x = 18. L. spicata var. prolifera, recorded as triploid 2n = 54, consisted of 30 metacentric chromosomes and 24 submetacentric chromosomes. Only one chromosome of the 11th group had a secondary constriction with a satellite in the short arm. L. Spicata was tetraploid 2n = 72 and consisted of four sets of 6 submetacentric chromosomes and 12 metacentric chromosomes without visible satellites. This paper provides further available data on Liriope chromosomes, and also indicates that L. spicata var. prolifera and L. spicata are probably separate species.     

The incidence of mistletoe (<Emphasis Type=Italic>Viscum album</Emphasis> ssp. <Emphasis Type=Italic>abietis</Emphasis>) on silver fir (<Emphasis Type=Italic>Abies alba</Emphasis>) in Croatia

The research on incidence of mistletoe (Viscum album ssp. abietis) on silver fir (Abies alba) was carried out in natural fir stands in Croatia. In the area of Gorski Kotar 32.8 % of the examined dominant fir trees were infected with mistletoe. The mistletoe incidence was presented according to the damage degrees of silver fir from 0 (healthy trees) to 4 (dead trees), rated by the crown defoliation and needle discoloration. With the increase of incidence, mistletoe spreads more onto more vital, i.e. less damaged trees. In the Dinaric Alps 28.6% of the examined fir trees were infected, this percentage amounting to 27.1% for the mountainous regions between the Sava and Drava rivers. The site and stand parameters (exposure, elevation, site quality, forest community and crown closure) were analysed in order to establish whether there was a correlation between these parameters and mistletoe incidence. There was a negative correlation between the elevation and mistletoe incidence. Among the compartments with a closed stands there were considerably less compartments with higher mistletoe incidence than among compartments with a sparsely closed and understocked stands. The other analysed site and stand parameters individually had no significant influence on mistletoe incidence. The correlation of silver fir mortality in 2004 and mistletoe incidence in 2002/03 was analysed, and their strong correlation was established. Mistletoe could be considered as a bioindicator of silver fir decline, and probably a significant contributor to that decline. In the areas where mistletoe incidence is great it can be presumed that silver fir is significantly damaged.     

Isolation and characterization of microsatellite markers for <Emphasis Type=Italic>Cedrela odorata</Emphasis> L. (Meliaceae), a high value neotropical tree

We describe 9 primers for amplification of microsatellite loci for the Neotropical tree Cedrela odorata L. (Meliaceae). Loci were isolated from an enriched library derived from a single DNA sample from a tree in Costa Rica. Levels of polymorphism were determined using samples from a large progeny trial. Across loci, the number of alleles ranged from 14 to 30. Observed heterozygosity levels ranged from 0.61 to 0.88. No linkage disequilibria were detected although some departures from Hardy-Weinberg equilibrium (HWE) were found, probably due to a Wahlund effect.