Phenotypic, genetic and genomic consequences of natural and synthetic polyploidization of Nicotiana attenuata and Nicotiana obtusifolia

Background and Methods

Polyploidy results in genetic turmoil, much of which is associated with new phenotypes that result in speciation. Five independent lines of synthetic allotetraploid N. × obtusiata (N × o) were created from crosses between the diploid N. attenuata (Na) (♂) and N. obtusifolia (No) (♀) and the autotetraploids of Na (NaT) and No (NoT) were synthesized. Their genetic, genomic and phenotypic changes were then compared with those of the parental diploid species (Na and No) as well as to the natural allotetraploids, N. quadrivalvis (Nq) and N. clevelandii (Nc), which formed 1 million years ago from crosses between ancient Na and No.

Key Results

DNA fingerprinting profiles (by UP-PCR) revealed that the five N × o lines shared similar but not identical profiles. Both synthetic and natural polyploidy showed a dosage effect on genome size (as measured in seeds); however, only Nq was associated with a genome upsizing. Phenotypic analysis revealed that at the cellular level, N × o lines had phenotypes intermediate of the parental phenotypes. Both allo- and autotetraploidization had a dosage effect on seed and dry biomass (except for NaT), but not on stalk height at first flower. Nc showed paternal (Na) cellular phenotypes but inherited maternal (No) biomass and seed mass, whereas Nq showed maternal (No) cellular phenotypes but inherited paternal (Na) biomass and seed mass patterns. Principal component analysis grouped Nq with N × o lines, due to similar seed mass, stalk height and genome size. These traits separated Nc, No and Na from Nq and N × o lines, whereas biomass distinguished Na from N × o and Nq lines, and NaT clustered closer to Nq and N × o lines than to Na.

Conclusions

Both allo- and autotetraploidy induce considerable morphological, genetic and genomic changes, many of which are retained by at least one of the natural polyploids. It is proposed that both natural and synthetic polyploids are well suited for studying the evolution of adaptive responses.Key words: Nicotiana, polyploidy, genome size, DNA fingerprinting, phenotypic variation, Solanaceae     

Lateral Dispersal and Foraging Behavior of Entomopathogenic Nematodes in the Absence and Presence of Mobile and Non-Mobile Hosts

Entomopathogenic nematodes have been classified into cruisers (active searchers) and ambushers (sit and wait foragers). However, little is known about their dispersal and foraging behavior at population level in soil. We studied lateral dispersal of the ambush foraging Steinernema carpocapsae (ALL strain) and cruise foraging Heterorhabditis bacteriophora (GPS11 strain) from infected host cadavers in microcosms (0.05 m²) containing Wooster silt-loam soil (Oxyaquic fragiudalf) and vegetation in the presence or absence of non-mobile and mobile hosts. Results showed that the presence of a non-mobile host (Galleria mellonella larva in a wire mesh cage) enhanced H. bacteriophora dispersal for up to 24 hr compared with no-host treatment, but had no impact on S. carpocapsae dispersal. In contrast, presence of a mobile host (G. mellonella larvae) increased dispersal of S. carpocapsae compared with no host treatment, but had no effect on H. bacteriophora dispersal. Also H. bacteriophora was better at infecting non-mobile than mobile hosts released into the microcosms and S. carpocapsae was better at infecting mobile than non-mobile hosts, thus affirming the established cruiser-ambusher theory. However, results also revealed that a large proportion of infective juveniles (IJs) of both species stayed near (≤ 3.8 cm) the source cadaver (88-96% S. carpocapsae; 67–79% H. bacteriophora), and the proportion of IJs reaching the farthest distance (11.4 cm) was significantly higher for S. carpocapsae (1.4%) than H. bacteriophora (0.4%) in the presence of mobile hosts. S. carpocapsae also had higher average population displacement than H. bacteriophora in the presence of both the non-mobile (5.07 vs. 3.6 cm/day) and mobile (8.06 vs. 5.3 cm/day) hosts. We conclude that the two species differ in their dispersal and foraging behavior at the population level and this behavior is affected by both the presence and absence of hosts and by their mobility.     

Interspecific relationships and origins of Taxaceae and Cephalotaxaceae revealed by partitioned Bayesian analyses of chloroplast and nuclear DNA sequences

The precise delimitation of Taxaceae and Cephalotaxaceae is not totally resolved. Some contradicting taxonomic proposals have been published, which demonstrates the difficulties in establishing a natural classification of the families and especially in proposing a relevant treatment within the genera Taxus and Cephalotaxus. The aims of this study are to contribute to the phylogeny and specific delineation of the two conifer families on the basis of molecular data. A cladistic analysis of the sequences of five chloroplast (matK, rbcL, trnL, trnL-trnF spacer, and psbA-trnH spacer) and one nuclear (ITS) molecular markers was carried out, both individually and in combination, by distance, parsimony, likelihood, and Bayesian methods. The results confirm that the two families are monophyletic. In the genus Taxus, T. floridana is the first-branching taxon; T. brevifolia and T. globosa cluster together and are sister to T. baccata; the endemic T. yunnanensis clusters with T. wallichiana in subclade B and is only distantly related with the other four Taxus species in China (subclade A); T. fuana is closer to T. baccata than to other Taxus species. Torreya jackii and A. formosana are the first-branching species within Torreya and Amentotaxus, respectively. C. koreana and C. wilsoniana could be treated as two varieties of C. harringtonia. The ancestral distribution area of Taxaceae and Cephalotaxaceae is restricted either to Southwest China or Southeast China by DIVA analysis. The relaxed molecular clock indicates that the deepest divergences in Taxus go back to the late-Cretaceous. psbA-trnH, rbcL third codon position, and matK first codon position contributed most to the separation of taxa in Discriminant function analysis. Our results confirm, on a basis of multiple molecular markers and a complete sampling of basic species, the suggested monophyly of Taxaceae and Cephalotaxaceae and propose interspecific relationships within each group, with profound nomenclatural and taxonomic implications. Combination of partitioned Bayesian analysis and likelihood-based methods produced a more robust phylogenetic hypothesis for the two studied families.     

Synthesis and degradation of termination and premature-termination fragments of beta-galactosidase in vitro and in vivo.

A number of abnormal polypeptides which are products of the lacZ gene in Escherichia coli have been characterized. A variety of experiments indicates that one class of at least nine different fragments arises from premature termination of protein synthesis. They have been detected as products of protein synthesis both in vitro and in vivo, although the percentage of fragments made in vitro is greater than that in vivo. The percentage of total Z gene-encoded protein which is found as fragments is estimated to be roughly 23% in vivo. This means that about 31% of the total number of β-galactosidase monomers synthesized are prematurely terminated molecules. The average molecular weight of the polypeptides produced is 91,000. This corresponds to a termination event occurring on the average once every 3200 codons. The sites at which termination occurs appear to be specific and are located primarily near the 3′-end of the gene.Polypeptides synthesized in vitro and in vivo from templates containing mutations in the Z gene have also been compared. Most of the mutationally generated fragments synthesized in vitro are stable, unlike their in vivo counterparts, which are often rapidly degraded. One fragment, however, generated by an early amber mutation in the Z gene, is degraded in the in vitro system. The mechanism of degradation appears to be specific for small abnormal polypeptides. Internal reinitiation polypeptides generated by nonsense mutations, which have been found in vivo, are not detected in the in vitro protein synthesis system under the conditions used here.     

A new species and synonymy of the Neotropical Eucelatoria Townsend and redescription of Myiodoriops Townsend

The New World tropics represents the most diverse region for tachinid parasitoids (Diptera: Tachinidae), but it also contains the most narrowly defined, and possibly the most confusing, tachinid genera of any biogeographic region. This over-splitting of genera and taxonomic confusion has limited progress toward our understanding the family in this region and much work is needed to revise, redefine, and make sense of the profusion of finely split taxa. In a recent analysis of the Neotropical genus Erythromelana Townsend, two species previously assigned to this genus, Euptilodegeeria obumbrata (Wulp) and Myiodoriops marginalis Townsend were reinstated as monotypic genera. In the present study, we demonstrate that Euptilodegeeria obumbrata (Wulp), previously assigned to three different genera, represents in fact a species of the large New World genus Eucelatoria Townsend, in which females possess a sharp piercer for oviposition. We also show that the species Eucelatoria carinata (Townsend) belongs to the same species group as Eucelatoria obumbrata, which we here define and characterize as the Eucelatoria obumbrata species group. Additionally, we describe Eucelatoria flava sp. n. as a new species within the Eucelatoria obumbrata species group. Finally, we redescribe the genus Myiodoriops Townsend and the single species Myiodoriops marginalis Townsend.     

Fatty acids and sterols of selected hyphochytriomycetes and chytridiomycetes

The fatty acids and sterols of eight Chytridiomycetes and two Hyphochytriomycetes, and fatty acids of the OomycetePythium gracile, were analyzed by gas-liquid chromatography. In addition to the fatty acids anticipated for fungi, the two Hyphochytriomycetes (Hyphochytrium catenoides andRhizidiomyces apophysatus) and four of the Chytridiomycetes (Catenaria anguillulae, Blastocladiella emersonii, Monoblepharella sp., andAllomyces macrogynus) contained arachidonic acid as a major fatty acid of the polar lipid fraction, and this fatty acid was detected as a minor component ofRhizophlyctis rosea andSpizellomyces punctatum. Eicosapentaenoic acid constituted 4.6% of the polar lipid fatty acids inMonoblepharella sp., and trace amounts were detected in several other species. Both the gamma (ω-6) and alpha (ω-3) isomers of linolenic acid were detected in all of the species analyzed. Cholesterol was the predominant (≥73%) sterol ofB. emersonii, R. rosea, A. macrogynus, andChytridium confervae, and a minor (<12%) component ofC. anguillulae, andH. catenoides. The major sterols of the other species included lanosterol (C. anguillulae, 45%), stigmasta-5,22-dien-3β-ol (H. catenoides, 51%), 24-ethyl-cholesterol (S. punctatum, 38%;H. catenoides, 17%;Monoblepharella sp., 70%; andR. apophysatus, 84%), 24-methyl-cholesterol (H. catenoides, 23%;R. apophysatus, 14%;S. punctatum, 53%), and 24-methylene cholesterol (Rhizophydium sphaerotheca, 51%). Neither ergosterol nor fucosterol was detected in any of the species studied.     

Impact of Transport Crate Reuse and of Catching and Processing on Campylobacter and Salmonella Contamination of Broiler Chickens

The influence of transport, catching, and processing on contamination of broiler chickens with Salmonella and Campylobacter was investigated. Transport crates were reused with high frequency and were often still contaminated with Salmonella and Campylobacter when they arrived at the farm despite the fact that they were washed at the factory, and thus they were a potential route of infection. These organisms contaminated the feathers of previously Campylobacter- and Salmonella-negative birds going to the processing plant and were isolated from processed carcasses, albeit at a low frequency. The Campylobacter types which were the predominant organisms on the live birds when they arrived at the processing plant were not necessarily the types that were most frequently isolated from processed carcasses. This finding may reflect cross-contamination that occurred during processing or differences in the tolerance of the strains to the hostile environments that the bacteria experienced. The process of catching and putting the birds in crates significantly increased the chance of contamination with Campylobacter (P < 0.001).     

Parallelismic homoplasy of leaf and stipule phenotypes among genetic variants of Pisum sativum and Medicago truncatula and some taxa of Papilionoideae, Caesalpinioideae and Mimosoideae subfamilies of the Leguminosae flora of Delhi

The leguminous flora of Delhi comprises 78 Papilionoideae, 24 Caesalpinioideae and 24 Mimosoideae species; 80 of them are perennials. Five types of imparipinnate and two types of paripinnate compound leaves were observed in the species. The paripinnate leaves are bipinnate in 25 species (mostly mimosoid) and bifoliate in two species. The imparipinnate leaves were trifoliate or multifoliate in 59 papilionoid species and multifoliate in 16 caesalpinioid species; four of the papilionoid species produced leafletted and tendrilled unipinnate leaves. Leaves were bifacially simple in 22 species, simple with ectopic terminal growth in one species and simple tendril in one species. Twenty-one species (mostly mimosoid) were devoid of stipules. In 82 species stipules were small and free. Stipules were large and lobed in 17 species and large and adnate in four species. Two species of Caesalpinioideae produce compound leaf-like stipules. All four stipule phenotypes of 126 species corresponded with stipular phenotypes observed in wild type, coch, st and coch st genotypes of the model legume P. sativum. The seven leaf phenotypes observed in 126 species corresponded with phenotypes expected among combinations of uni (uni-tac), af, ins, mfp and tl mutants of P. sativum and sgl1, cfl1, slm1 and palm1 mutants of M. truncatula, also an IRL model legume. All the variation in leaf and stipule morphologies observed in the leguminous flora of Delhi could be explained in terms of the gene regulatory networks already revealed in P. sativum and M. truncatula. It is hypothesized that the ancestral gene regulatory networks for leaves and stipules produced in Leguminosae were like that prevalent in P. sativum.     

Structural polypeptides and products of late genes of bacteriophage Mu: Characterization and functional aspects

This paper describes the identification and functional role of late gene products of bacteriophage Mu, including an analysis of the structural proteins of the Mu virion.In vitro reconstitution of infectious phage particles has shown that four genes (E, D, I, J) control the formation of phage heads and that a cluster of eight genes (K, L, M, N, P, Q, R, S) controls the formation of phage tails.Sodium dodecyl sulphate/polyacrylamide gel electrophoresis of Mu polypeptides synthesized in Escherichia coli minicells infected by Mu phages carrying amber mutations in various late genes has resulted in the identification of the products of gene C (15.5 × 10³M_r); H (64 × 10³M_r); F (54 × 10³M_r); G (16 × 10³M_r); L (55 × 10³M_r); N (60 × 10³M_r); P (43 × 10³M_r) and S (56 × 10³M_r). Minicells infected with λpMu hybrid phages and deletion mutants of Mu were used to identify polypeptides encoded by the V-β region of the Mu genome. These are the products of genes V, W or R (41.5 × 10³M_r, and 45 × 10³M_r); U (20.5 × 10³M_r) and of genes located in the β region (24 × 10³M_r (gpgin) and 37 × 10³M_r (possibly gpmom)).Analytical separation of the proteins of the Mu virion revealed that it consists of a major head polypeptide with a molecular weight of 33 × 10³, a second head polypeptide of 54 × 10³ (gpF) and two major tail polypeptides with molecular weights of 55 × 10³ and 12.5 × 10³ (gpL and gpY, respectively). In addition, there are five minor components of the tail (including gpN, gpS and gpU) and approximately seven minor components of the head structure of the virion (including gpH).     

Growth and Reproduction of Glyphosate-Resistant and Susceptible Populations of Kochia scoparia

Evolution of glyphosate-resistant kochia is a threat to no-till wheat-fallow and glyphosate-resistant (GR) cropping systems of the US Great Plains. The EPSPS (5-enol-pyruvylshikimate-3-phosphate synthase) gene amplification confers glyphosate resistance in the tested Kochia scoparia (L.) Schrad populations from Montana. Experiments were conducted in spring to fall 2014 (run 1) and summer 2014 to spring 2015 (run 2) to investigate the growth and reproductive traits of the GR vs. glyphosate-susceptible (SUS) populations of K. scoparia and to determine the relationship of EPSPS gene amplification with the level of glyphosate resistance. GR K. scoparia inbred lines (CHES01 and JOP01) exhibited 2 to 14 relative copies of the EPSPS gene compared with the SUS inbred line with only one copy. In the absence of glyphosate, no differences in growth and reproductive parameters were evident between the tested GR and SUS inbred lines, across an intraspecific competition gradient (1 to 170 plants m^-2). GR K. scoparia plants with 2 to 4 copies of the EPSPS gene survived the field-use rate (870 g ha^-1) of glyphosate, but failed to survive the 4,350 g ha^-1 rate of glyphosate (five-times the field-use rate). In contrast, GR plants with 5 to 14 EPSPS gene copies survived the 4,350 g ha^-1 of glyphosate. The results from this research indicate that GR K. scoparia with 5 or more EPSPS gene copies will most likely persist in field populations, irrespective of glyphosate selection pressure.     

Biodiversity and physiological characteristics of Antarctic and Arctic lichens-associated bacteria

The diversity and physiological characteristics of culturable bacteria associated with lichens from different habitats of the Arctic and Antarctica were investigated. The 68 retrieved isolates could be grouped on the basis of their 16S rRNA gene sequences into 26 phylotypes affiliated with the phyla Actinobacteria, Bacteroidetes, Deinococcus-Thermus, and Firmicutes and with the classes Alphaproteobacteria, Betaproteobacteria, and Gammaproteobacteria. Isolates belonging to the Alphaproteobacteria were the most abundant, followed by those belonging to Actinobacteria, Betaproteobacteria, Gammaproteobacteria, Bacteroidetes, Firmicutes, and Deinococcus-Thermus. Phylogenetic analysis showed that approximately 21 % of the total isolates represented a potentially novel species or genus (≤97 % sequence similarity). Strains belonging to the genera Sphingomonas, Frondihabitans, Hymenobacter, and Burkholderia were recovered from lichen samples from both geographic locations, implying common and important bacterial functions within lichens. Extracellular protease activities were detected in six isolates, affiliated with Burkholderia, Frondihabitans, Hymenobacter, Pseudomonas, and Rhodanobacter. Extracellular lipase activities were detected in 37 isolates of the genera Burkholderia, Deinococcus, Frondihabitans, Pseudomonas, Rhodanobacter, Sphingomonas, and Subtercola. This is the first report on the culturable bacterial diversity present within lichens from Arctic and Antarctica and the isolates described herein are valuable resources to decode the functional and ecological roles of bacteria within lichens. In addition, the low similarity (≤97 %) of the recovered isolates to known species and their production of cold-active enzymes together suggest that lichens are noteworthy sources of novel bacterial strains for use in biotechnological applications.     

Discosorids and Oncocerids (Cephalopoda) of the Middle Ordovician Kunda and Aseri Regional Stages of Baltoscandia and the early evolution of these groups

Discosorid and oncocerid cephalopods of the Kunda and Aseri Regional Stage (Darriwilian, Middle Ordovician) from Öland Island, Sweden and North Estonia are described for the first time. We demonstrate that a high generic and specific diversity of Oncocerida was already established in the earliest Darriwilian. The new oncocerid species Neumatoceras borense nov. sp., N. breviborense nov. sp., Paramiamoceras breviventrum nov. sp., Richardsonoceras gastroscopium nov. sp., R. gerhardi nov. sp., R. goldmanni nov. sp., R. haelluddenense nov. sp., Richardsonoceroides kingi nov. sp., R. oelandense nov. sp., and R. rhytium nov. sp. are erected; Richardsonoceroides schiefferdeckeri (Dewitz) is revised. The new graciloceratid Kundoceras evansi nov. gen., nov. sp. is erected. A review of records of early oncocerids from China shows that Richardsonoceras Foerste is probably the most primitive oncocerid and supports the hypothesis that oncocerids evolved in the early Floian or latest Tremadocian from a Bassleroceras Ulrich and Foerste-like ancestor. The new discosorid genus Paldoceras nov. gen. and species Paldoceras paldiskense nov. sp. and P. neptunsakerense nov. sp. are erected. These discosorids record the morphological link between the late Darriwilian Ruedemannoceras Flower and the early Floian Apocrinoceras Teichert and Glenister, which is considered to be the earliest discosorid.     

Molecular evolution and sequence divergence of plant chalcone synthase and chalcone synthase-Like genes

Plant chalcone synthase (CHS) and CHS-Like (CHSL) proteins are polyketide synthases. In this study, we evaluated the molecular evolution of this gene family using representative types of CHSL genes, including stilbene synthase (STS), 2-pyrone synthase (2-PS), bibenzyl synthase (BBS), acridone synthase (ACS), biphenyl synthase (BIS), benzalacetone synthase, coumaroyl triacetic acid synthase (CTAS), and benzophenone synthase (BPS), along with their CHS homologs from the same species of both angiosperms and gymnosperms. A cDNA-based phylogeny indicated that CHSLs had diverse evolutionary patterns. STS, ACS, and 2-PS clustered with CHSs from the same species (late diverged pattern), while CTAS, BBS, BPS, and BIS were distant from their CHS homologs (early diverged pattern). The amino-acid phylogeny suggested that CHS and CHSL proteins formed clades according to enzyme function. The CHSs and CHSLs from Polygonaceae and Arachis had unique evolutionary histories. Synonymous mutation rates were lower in late diverged CHSLs than in early diverged ones, indicating that gene duplications occurred more recently in late diverged CHSLs than in early diverged ones. Relative rate tests proved that late diverged CHSLs had unequal rates to CHSs from the same species when using fatty acid synthase, which evolved from the common ancestor with the CHS superfamily, as the outgroup, while the early diverged lineages had equal rates. This indicated that late diverged CHSLs experienced more frequent mutation than early diverged CHSLs after gene duplication, allowing obtaining new functions in relatively short period of time.     

Phylogeny of 16S rRNA and nifH genes and regulation of nitrogenase activity by oxygen and ammonium in the genus Paenibacillus

All Paenibacillus 16S rDNA sequences, except for that of Paenibacillus massiliensis T7, formed a coherent cluster, distinct from gram-positive nitrogen-fixing Clostridium pasteurianum and Heliobacterium chlorum. All Paenibacillus NifH sequences formed two main clusters. Cluster I encompassing the NifH sequences from most of members of Paenibacillus spp., such as Paenibacillus azotofixans NifH1 and NifH2, Paenibacillus polymyxa and Paenibacillus macerans. Cluster II including only P. azotofixans NifH3. Curiously, three copies of nifH genes of Paenibacillus sabine T27 clustered within P. azotofixans cluster I (NifH1 and NifH2). The effect of O₂ and ammonium on nitrogenase activity was studied with 14 different nitrogenfixing Paenibacillus strains. The optimal oxygen concentration level for all Paenibacillus strains is in the 0 to 0.05% range, similar to that for Klebsiella pneumoniae. In all Paenibacillus strains, the highest nitrogenase activity is obtained in the condition of 0?C0.1 mM NH₄Cl and the increase of NH₄Cl from 0.1 to 5 mM caused a rapid inhibition of nitrogenase activity. However, the inhibition was reversible in the presence of 200 mM NH₄Cl in some Paenibacillus strains. It is the first time to use almost all of the recognized nitrogen-fixing Paenibacilus spp. to investigate the phylogeny of 16S rRNA and nifH genes. The data that the inhibition of O₂ and ammonium on nitrogenase acitivity will provide a base for studying the molecular regulatory mechanism of nitrogen fixation in the genus Paenibacillus.     

Effects of dietary inclusion of citrus pulp and rockrose soft stems and leaves on lamb meat quality and fatty acid composition

Meat from lambs finished with high-starch diets often contains low concentration of vaccenic (t11-18:1) and rumenic (c9,t11-18:2) acids and high concentration of t10-18:1. We hypothesized that replacing cereals by dehydrated citrus pulp (DCP) and the inclusion of tanniferous feed sources in oil supplemented diets might reduce the accumulation of t10-18:1 and increase the t11-18:1 and c9,t11-18:2 in lamb meat, without affecting the productive performance. In total, 32 lambs were assigned to four diets which combine two factors: basal diet (BD) (cereals v. DCP) and Cistus ladanifer (CL) (0 v. 150 g/kg dry matter). Feed intake, average daily weight gain and carcass traits were not affected by treatments, except for dressing percentage that was reduced with DCP (P=0.046). Both DCP and C. ladanifer reduced tenderness and juiciness of meat, and C. ladanifer also reduced (P<0.001) meat overall acceptability. Intramuscular fat and the concentration of saturated and polyunsaturated fatty acids (FA) were not affected (P>0.05) by diets. However, DCP increased the proportions of odd-chain FA (P=0.005) and several minor biohydrogenation (BH) intermediates in meat lipids. C. ladanifer had few effects on meat FA profile. The proportions of t11-18:1 and c9,t11-18:2 were high in all diets (5.4% and 1.5% of total FA, respectively) and were not influenced by the treatments. Basal diet and CL showed some significant interactions concerning FA composition of intramuscular fat. In diets without C. ladanifer, replacement of cereals by DCP increased the 18:0 (P<0.05) and decreased t10,c12-18:2 (P<0.05), t10-18:1 (P<0.10) and t10-/t11-18:1 ratio (P<0.10) with a large reduction of the individual variation for t10-18:1 and of t10-/t11-18:1 ratio. Combined with cereals, C. ladanifer increased 18:0 and reduced the BH intermediates in meat. Replacement of cereals by DCP seems to promote a more predictable FA profile in lamb meat, reducing the risk of t10-shifted BH pathways in the rumen.     

Synthesis of azide and amide analogs of platelet-activating factor and related derivatives

2-Azido-2-deoxy-1-O-hexadecyl-sn-glycero-3-phosphorylcholine was prepared in good yield from D-mannitol via 3-O-hexadecyl-2-O-methanesulfonyl-1-O-triphenylmethyl-sn-glycerol. Nucleophilic displacement of the 2-methanesulfonate function by benzoate or azide ion proceeded with inversion of configuration (Sn2) without racemization. Hydrogenation of the azidophospholipid gave 2-amino-2-deoxy-1-O-hexadecyl-sn-glycero-3-phosphorylcholine which is a versatile intermediate for the preparation of amide analogs of platelet-activating factor and related derivatives. The synthesis of 2-deoxy-2-fluoro-1-O-hexadecyl-sn-glycero-3-phosphorylcholine was also described.     

Identification and Pathogenic Potential of Clinical Bacillus and Paenibacillus Isolates

The soil-related Bacillus and Paenibacillus species have increasingly been implicated in various human diseases. Nevertheless, their identification still poses problems in the clinical microbiology laboratory and, with the exception of Bacillus anthracis and Bacillus cereus, little is known on their pathogenicity for humans. In this study, we evaluated the use of matrix-assisted laser desorption—ionization time of flight mass spectrometry (MALDI-TOF MS) in the identification of clinical isolates of these genera and conducted genotypic and phenotypic analyses to highlight specific virulence properties. Seventy-five clinical isolates were subjected to biochemical and MALDI-TOF MS identification. 16S rDNA sequencing and supplemental tests were used to solve any discrepancies or failures in the identification results. MALDI-TOF MS significantly outperformed classical biochemical testing for correct species identification and no misidentification was obtained. One third of the collected strains belonged to the B. cereus species, but also Bacillus pumilus and Bacillus subtilis were isolated at high rate. Antimicrobial susceptibility testing showed that all the B. cereus, B. licheniformis, B. simplex, B. mycoides, Paenibacillus glucanolyticus and Paenibacillus lautus isolates are resistant to penicillin. The evaluation of toxin/enzyme secretion, toxin-encoding genes, motility, and biofilm formation revealed that B. cereus displays the highest virulence potential. However, although generally considered nonpathogenic, most of the other species were shown to swim, swarm, produce biofilms, and secrete proteases that can have a role in bacterial virulence. In conclusion, MALDI-TOF MS appears useful for fast and accurate identification of Bacillus and Paenibacillus strains whose virulence properties make them of increasing clinical relevance.     

Preparation and standardization of antigens of Histoplasma capsulatum and Coccidioides immitis

Methods have been developed for the separation and purification of various antigenically active cellular components ofH. capsulatum (34) andC. immitis (35), and chemical procedures have been employed in the characterization of these antigens. The concluding remarks illustrate the current trends in immunological studies ofH. capsulatum andC. immitis. Hopefully, these approaches will provide knowledge and insight into the basic biological properties ofH. capsulatum andC. immitis and should, in the future, culminate in the physicochemical characterization of their important antigens.     

The effects of temperature and aging of Drosophila males on the frequencies of XXY and XO progeny

Drosophila males of the constitution y/y⁺Y were aged for 10 days at 25° and 10° and then mated daily for 13 days at 25° to virgin yw (XX) females. The total frequencies of exceptional XXY and XO offspring to which the father contributed either (a) both an X and a Y chromosome, or (b) neither of them, were highly significantly more numbers in the 7th- and 8th-day broods of the 10° than the 25° pre-aged series. In all experiments the frequency of paternal XO exceptions was greatly in excess of that of XXY exceptions. The data on brood patterns suggest that the stages most sensitive to the production of paternal exceptions by pre-aging at 10° are those of the primary spermatocytes. The same stages are also sensitive to low temperature induction of temporarily low fertility.     

Karyomorphology and relationships of Celtidaceae and Ulmaceae (Urticales)

Based on karyomorphological features, six (examined in this study) of nine genera of Celtidaceae are divided into three groups: 1)Celtis, Parasponia, Pteroceltis andTrema; 2)Aphananthe; 3)Gironniera, and six genera of Ulmaceae into two: 1)Holoptelea andPhyllostylon; 2)Hemiptelea, Planera, Ulmus andZelkova. The first four genera share the simple chromocenter type at the resting stage andx-10, with all chromosomes with submedian or median centromeres (frequency of chromosomes with subterminal or terminal centromeres 0%, although uncertain inTrema).Aphananthe hasx=13, but resembles the above four genera in other features.Gironniera is distinct from all other Celtidaceae in having the diffuse-complex chromocenter type andx=14, features which occur in Ulmaceae. InGironniera the frequency of chromosomes with subterminal or terminal centromeres is 43%, a proportion similar to those inHoloptelea (36%) andPhyllostylon (58%) of Ulmaceae. All six genera of Ulmaceae havex=14, yetHemiptelea, Planera, Ulmus andZelkova are distinct fromHoloptelea andPhyllostylon (with the simple chromocenter type) in having the diffuse-complex chromocenter type and in predominantly possessing chromosomes with subterminal or terminal centromeres (93%). Evidence from karyomorphology, as well as from other sources, suggests 1) thatAphananthe (x=13) is most distantly related to all genera withx=10 within Celtidaceae, 2) thatGironniera may have a key role for understanding evolutionary relationships between Celtidaceae and Ulmaceae, and 3) thatHoloptelea andPhyllostylon represent derivatives of a line that diverged early from a common ancestor or all Ulmaceae. On the basis of comparisons with other Urticales and the putative outgroups of the order, it is also suggested that the chromosome morphology of Ulmaceae represents the more derived state in Urticales.