Evolutionary divergence of the paralogs Methoprene tolerant (Met) and germ cell expressed (gce) within the genus Drosophila

Juvenile hormone (JH) signaling underpins both regulatory and developmental pathways in insects. However, the JH receptor is poorly understood. Methoprene tolerant (Met) and germ cell expressed (gce) have been implicated in JH signaling in Drosophila. We investigated the evolution of Met and gce across 12 Drosophila species and found that these paralogs are conserved across at least 63 million years of dipteran evolution. Distinct patterns of selection found using estimates of dN/dS ratios across Drosophila Met and gce coding sequences, along with their incongruent temporal expression profiles in embryonic Drosophila melanogaster, illustrate avenues through which these genes have diverged within the Diptera. Additionally, we demonstrate that the annotated gene CG15032 is the 5′ terminus of gce.In mosquitoes and beetles, a single Met-like homolog displays structural similarity to both Met and gce, and the intron locations are conserved with those of gce. We found that Tribolium and mosquito Met orthologs are assembled from Met- and gce-specific domains in a modular fashion. Our results suggest that Drosophila Met and gce experienced divergent evolutionary pressures following the duplication of an ancestral gce-like gene found in less derived holometabolous insects.     

Phyllocarida and the origin of the Malacostraca

Tentative homologies with Decapoda are proposed for grooves on the carapace of Echinocaris and Montecaris. Together with F.R. Schram's hoploid trend, reinforced by the discovery of a raptorial limb in Sairocaris, this may strengthen a phyllocarid origin for Eumalacostraca. Both these trends, however, may be parallelisms, and phylogenetic analysis is required. Pleopods are here proved to have been present in additional Archaeostraca, which improves their ancestral condition. E. Dahl's view of the phyllocarids as an evolutionary dead end is falsified from fossils, and the phyllocarids are supported as a stem group of the Malacostraca.     

Rhizobium pisi sv. trifolii K3.22 harboring nod genes of the Rhizobium leguminosarum sv. trifolii cluster

The taxonomic status of the Rhizobium sp. K3.22 clover nodule isolate was studied by multilocus sequence analysis (MLSA) of 16S rRNA and six housekeeping chromosomal genes, as well as by a subsequent phylogenic analysis. The results revealed full congruence with the Rhizobium pisi DSM 30132^T core genes, thus supporting the same taxonomic position for both strains. However, the K3.22 plasmid symbiosis nod genes demonstrated high sequence similarity to Rhizobium leguminosarum sv. trifolii, whereas the R. pisi DSM 30132^Tnod genes were most similar to R. leguminosarum sv. viciae. The strains differed in the host range nodulation specificity, since strain K3.22 effectively nodulated red and white clover but not vetch, in contrast to R. pisi DSM 30132^T, which effectively nodulated vetch but was not able to nodulate clover. Both strains had the ability to form nodules on pea and bean but they differed in bean cultivar specificity. The R. pisi K3.22 and DSM 30132^T strains might provide evidence for the transfer of R. leguminosarum sv. trifolii and sv. viciae symbiotic plasmids occurring in natural soil populations.     

Systematics and evolution of the genus Torrubiella (Hypocreales, Ascomycota)

Torrubiella is a genus of arthropod-pathogenic fungi that primarily attacks spiders and scale insects. Based on the morphology of the perithecia, asci, and ascospores, it is classified in Clavicipitaceae s. lat. (Hypocreales), and is considered a close relative of Cordyceps s. 1., which was recently reclassified into three families (Clavicipitaceae s. str., Cordycipitaceae, Ophiocordycipitaceae) and four genera (Cordyceps s. str, Elaphocordyceps, Metacordyceps, and Ophiocordyceps). Torrubiella is distinguished morphologically from Cordyceps s. lat. mainly by the production of superficial perithecia and the absence of a well-developed stipitate stroma. To test and refine evolutionary hypotheses regarding the placement of Torrubiella and its relationship to Cordyceps s. lat., a multi-gene phylogeny was constructed by conducting ML and Bayesian analyses. The monophyly of Torrubiella was rejected by these analyses with species of the genus present in Clavicipitaceae, Cordycipitaceae, and Ophiocordycipitaceae, and often intermixed among species of Cordyceps s. lat. The morphological characters traditionally used to define the genus are, therefore, not phylogenetically informative, with the stipitate stromata being gained and/or lost several times among clavicipitaceous fungi. Two new genera (Conoideocrella, Orbiocrella) are proposed to accommodate two separate lineages of torrubielloid fungi in the Clavicipitaceae s. str. In addition, one species is reclassified in Cordyceps s. str. and three are reclassified in Ophiocordyceps. The phylogenetic importance of anamorphic genera, host affiliation, and stipitate stromata is discussed.     

Mycobiota associated with the ambrosia beetle Scolytodes unipunctatus (Coleoptera: Curculionidae, Scolytinae)

Ambrosia beetles (Coleoptera: Scolytinae) are associated with strictly entomochoric and mutualistic fungi. We studied the mycobiota associated with Scolytodes unipunctatus, ambrosia beetles that infest Cecropia trees in Central America. Isolates were characterized using morphology and rDNA sequences (ITS region, LSU, and SSU rDNA). Four species are described here: Raffaelea scolytodis sp. nov. (Ophiostomatales), Gondwanamyces scolytodis sp. nov., Custingophora cecropiae sp. nov., and Graphium sp. (Microascales). The genus Custingophora is emended to include Knoxdaviesia anamorphs of Gondwanamyces based on uniformity of DNA sequences and phenotype.     

Expression profiling of the triterpene saponin biosynthesis genes FPS, SS, SE, and DS in the medicinal plant Panax notoginseng

Panax notoginseng (Burk) F. H. Chen, an economically significant medicinal plant with hemostatic and health tonic activities, has been used in Traditional Chinese Medicine (TCM) for more than 3000 years. Triterpene saponins are responsible for most of the pharmacological activities of P. notoginseng. Here, we cloned five cDNA sequences encoding the key enzymes involved in triterpene saponin biosynthesis, namely, PnFPS, PnSS, PnSE1, PnSE2, and PnDS, and analyzed the conserved domains and phylogenetics of their corresponding proteins. Their organ-specific expression patterns in four-year-old P. notoginseng were detected by real-time PCR, showing that they were all most highly expressed in flowers. In addition, four of the genes, excluding PnSE2, were upregulated in leaves following stimulation with methyl jasmonate. This study is the first comprehensive analysis of the expression patterns of pivotal genes for triterpene saponin biosynthesis in P. notoginseng and provides a basis to further elucidate the molecular mechanism for the biosynthesis of these medically important compounds.     

Interconversion of trans, trans and cis, trans farnesol by enzymes from Andrographis

When trans, trans-farnesol [4,8,12-¹⁴C₃,1-³H₂] is isomerized to cis, trans-farnesol by soluble enzymes from Andrographis paniculata tissue cultures, 50% of the tritium label is lost. The same loss is observed when isomerization occurs in the opposite direction. This is in accordance with the proposed mechanism for isomerization via aldehydes.     

On the roles of Notch, Delta, kuzbanian, and inscuteable during the development of Drosophila embryonic neuroblast lineages

The generation of cellular diversity in the nervous system involves the mechanism of asymmetric cell division. Besides an array of molecules, including the Par protein cassette, a heterotrimeric G protein signalling complex, Inscuteable plays a major role in controlling asymmetric cell division, which ultimately leads to differential activation of the Notch signalling pathway and correct specification of the two daughter cells. In this context, Notch is required to be active in one sibling and inactive in the other. Here, we investigated the requirement of genes previously known to play key roles in sibling cell fate specification such as members of the Notch signalling pathway, e.g., Notch (N), Delta (Dl), and kuzbanian (kuz) and a crucial regulator of asymmetric cell division, inscuteable (insc) throughout lineage progression of 4 neuroblasts (NB1-1, MP2, NB4-2, and NB7-1). Notch-mediated cell fate specification defects were cell-autonomous and were observed in all neuroblast lineages even in cells born from late ganglion mother cells (GMC) within the lineages. We also show that Dl functions non-autonomously during NB lineage progression and clonal cells do not require Dl from within the clone. This suggests that within a NB lineage Dl is dispensable for sibling cell fate specification. Furthermore, we provide evidence that kuz is involved in sibling cell fate specification in the central nervous system. It is cell-autonomously required in the same postmitotic cells which also depend on Notch function. This indicates that KUZ is required to facilitate a functional Notch signal in the Notch-dependent cell for correct cell fate specification. Finally, we show that three neuroblast lineages (NB1-1, NB4-2, and NB7-1) require insc function for sibling cell fate specification in cells born from early GMCs whereas insc is not required in cells born from later GMCs of the same lineages. Thus, there is differential requirement for insc for cell fate specification depending on the stage of lineage progression of NBs.     

Genetic regulation of engrailed and wingless in Tribolium segmentation and the evolution of pair-rule segmentation

In Drosophila, primary pair-rule genes establish the parasegmental boundaries and indirectly control the periodic expression of the segment polarity genes engrailed (en) and wingless (wg) via regulation of secondary pair-rule genes. Although orthologs of some Drosophila pair-rule genes are not required for proper segmentation in Tribolium, segmental expression of Tc-en and Tc-wg is conserved. To understand how these segment polarity genes are regulated, we examined the results of expressing one or two pair-rule genes in the absence of the other known pair-rule genes. Expression of one or both of the secondary pair-rule genes, Tc-sloppy-paired (Tc-slp) and Tc-paired (Tc-prd), activated Tc-wg in the absence of the primary pair-rule genes, Tc-even-skipped (Tc-eve), Tc-runt (Tc-run) and Tc-odd-skipped (Tc-odd). Tc-eve alone failed to activate Tc-wg or Tc-en, but in combination with Tc-run or Tc-prd activated Tc-en. These results, interpreted within the pair-rule gene expression patterns, suggest separate models for the genetic regulation of the juxtaposed expression of Tc-wg and Tc-en at odd- and even-numbered parasegmental boundaries, respectively. Conserved interactions between eve and prd at the anterior boundary of odd-numbered parasegments may reflect an ancestral segmentation mechanism that functioned in every segment prior to the evolution of pair-rule segmentation.     

Recent advances in the cryopreservation of shoot-derived germplasm of economically important fruit trees of Actinidia, Diospyros, Malus, Olea, Prunus, Pyrus and Vitis

This paper presents the advances made over the last decade in cryopreservation of economically important vegetatively propagated fruit trees. Cryopreservation protocols have been established using both dormant buds sampled on field-grown plants and shoot tips sampled on in vitro plantlets. In the case of dormant buds, scions are partially dehydrated by storage at − 5 °C, and then cooled slowly to − 30 °C using low cooling rates (c.a. 1 °C/h) before immersion in liquid nitrogen. After slow rewarming and rehydration of samples, regrowth takes place either through grafting of buds on rootstocks or excision of apices and inoculation in vitro. In the case of shoot tips of in vitro plantlets, the cryopreservation techniques employed are the following: controlled rate cooling procedures involving slow prefreezing followed by immersion in liquid nitrogen or vitrification-based procedures including encapsulation–dehydration, vitrification, encapsulation–vitrification and droplet-vitrification. The current status of cryopreservation for a series of fruit tree species including Actinidia, Diospyros, Malus, Olea, Prunus, Pyrus and Vitis is presented. Routine application of cryopreservation for long-term germplasm storage in genebanks is currently limited to apple and pear, for which large cryopreserved collections have been established at NCGRP, Fort Collins (USA), using dormant buds and in vitro shoot tips, respectively. However, there are a growing number of examples of pilot scale testing experiments under way for different species in various countries. Progress in the further development and application of cryopreservation techniques will be made through a better understanding of the mechanisms involved in the induction of tolerance to dehydration and cryopreservation in frozen explants.     

Occurrence of Nosema oryzaephili in Cryptolestes ferrugineus and transfer to the genus Paranosema

A microsporidium that closely resembles Paranosema species at the level of the light microscope was isolated from the rusty grain beetle, Cryptolestes ferrugineus. It’s identity as Nosema oryzaephili (originally described from Oryzaephilus surinamensis) was confirmed by comparison with a known isolate of N. oryzaephili based on spore size, small subunit rDNA sequence, and relative infectivity to O. surinamensis, Tribolium castaneum, and Ephestia kuehniella. Phylogenetic analysis of the small subunit rDNA indicates clearly that this species belongs in the genus Paranosema and thus the designation Paranosema oryzaephili (Burges, Canning and Hurst) is proposed. In spite of the abundance, economic importance, and world-wide distribution of C. ferrugineus, this is the first report of a microsporidial infection in this species. This is also the first report of P. oryzaephili in the new world.     

Serological evidence confirming the assignment of phaseolus aureus and P. mungo to the genus vigna

Endopeptidase activity was detected in extracts of cotyledons of 11 species of Vigna and Phaseolus Antibodies against the purified protease isolated from the cotyledons of 5-day-old P.aureus seedlings inhibited the activity of that enzyme in crude extracts of cotyledons. A similar inhibition was obtained with P. mungo, P. adenanthus and 4 species of Vigna, while there was no inhibition of endopeptidase activity in extracts of cotyledons of 4 species of Phaseolus. Immunodiffusion tests proved that the protease of Vigna is distinct from that of Phaseolus. The evidence supports the reassignment of P. aureus and P. mungo to the genus Vigna and indicates that the names Vigna radiata and Vigna mungo are more appropriate than P. aureus and P. mungo for green gram and black gram respectively.     

Chemosystematics of section Palaeonarthex of the Genus Ferula

Ferula species of the section Palaeonarthex contains various sesquiterpenoids. While C-15 terpenoid coumarins of the iresan type are the most characteristic constituents of species in the subsection Foetidissimae, sesquiterpene lactones and C-10 terpenoid coumarns are found in the subsection Tumidae and esters of the sesquiterpene alcohols in the subsection Jaeschkaenae. The biogenetic relationships of the Ferula sesquiterpenoids and their chemosystematic value are discussed.     

Bioassay of Entomophthora against the spotted alfalfa aphid Therioaphis trifolii f. maculata

A method for the bioassay of Enthomophthora spp. against aphids is described. Twenty-four isolates comprising five species of fungi were screened for activity against Therioaphis trifolii f. maculata. The only isolates with a high level of activity were those of E. sphaerosperma obtained from aphids. The initial bioassays with E. sphaerosperma indicated that aphids, starved for 24 hr during inoculation with E. sphaerosperma primary spores, were less susceptible than those removed from the plants, for just the period of exposure to the primary spore shower. Using the latter procedure, five assays of the most pathogenic isolate gave a mean LC₅₀ of 11.3 primary spores/mm² while bioassays of four other isolates gave LC₅₀ values ranging from 15.7 to 27.3 spores/mm². The potential of E. sphaerosperma as a microbial control agent for T. trifolii f. maculata in Australia is discussed.