基于16S rRNA和rpoB基因的分子系统发育分析在铜绿假单胞菌鉴定中的应用

为对比16S rRNA和rpo B基因分子系统发育分析与传统表型分类法对铜绿假单胞菌的鉴定,评估16S rRNA和rpo B基因序列分析在铜绿假单胞菌鉴定中的应用,用表型分类方法对临床自动微生物鉴定系统鉴定为铜绿假单胞菌的23株分离株进行再鉴定,PCR扩增23株分离株16S rRNA和rpo B基因片段,并测序进行系统发育分析。结果表明,表型再鉴定结果与自动微生物鉴定系统鉴定结果一致。基于两个基因的系统发育分析均显示分离株p22与不动杆菌属序列聚为一枝,其余22株分离株与铜绿假单胞菌序列聚为一枝。因此p22应鉴定为不动杆菌,16S rRNA和rpo B基因序列分析均能准确鉴定铜绿假单胞菌并能较好建立假单胞菌属内种间关系。     

Genetic and phenotypic evidence for two groups of <Emphasis Type="Italic">Oenococcus oeni</Emphasis> strains and their prevalence during winemaking

Polymerase chain reaction (PCR)–denaturing gradient gel electrophoresis was the most relevant method to follow the diversity of lactic acid bacteria during winemaking. By targeting the rpoB gene, two types of Oenococcus oeni strains were distinguished resulting from a single mutation in the rpoB region targeted in PCR and generating two different electrophoresis profiles. The first one prevailed during fermentation and the second during ageing. Some strains of each type were isolated during winemaking and were studied using several genetic methods (real-time PCR, PCR-random amplified polymorphic DNA, multiple locus sequence typing and the presence of gene markers). Physiological characters related to environmental conditions were examined. The results confirmed the relevance of the rpoB mutation for characterising the two O. oeni subgroups. The relationship between the physiological response to stress and the rpoB genetic groups raised the question of O. oeni intraspecies grouping. A possible division within this species, of great technological interest to the wine industry, was also raised.     

The plastid rpoA gene encoding a protein homologous to the bacterial RNA polymerase alpha subunit is expressed in pea chloroplasts

Summary The gene rpoA, encoding a protein homologous to the alpha subunit of RNA polymerase from Escherichia coli has been located in pea chloroplast DNA downstream of the petD gene for subunit IV of the cytochrome b-f complex. Nucleotide sequence analysis has revealed that rpoA encodes a polypeptide of 334 amino acid residues with a molecular weight of 38916. Northern blot analysis has shown that rpoA is co-transcribed with the gene for ribosomal protein S11. A lacZ-rpoA gene-fusion has been constructed and expressed in E. coli. Antibodies raised against the fusion protein have been employed to demonstrate the synthesis of the rpoA gene product in isolated pea chloroplasts. Western blot analysis using these antibodies and antibodies against the RNA polymerase core enzyme from the cyanobacterium, Anabaena 7120, has revealed the presence of the gene product in a crude RNA polymerase preparation from pea chloroplasts.     

Nucleotide sequence of the maize chloroplast rpo B/C1/C2 operon: Comparison between the derived protein primary structures from various organisms with respect to functional domains

Summary The genes (rpo B/C₁/C₂) coding for the , , subnits of maize (Zea mays) chloroplast RNA polymerase have been located on the plastome and their nucleotide sequences established. The operon is part of a large inversion with respect to the tobacco and spinach chloroplast genomes and is flanked by the genes trnC and rps2. Notable features of the nucleotide sequence are the loss of an intron in rpoC₁, and an insertion of approximately 450 by in rpOC₂ compared to the dicotyledons tobacco, spinach and liver-wort. The derived amino acid sequence of this additional monocotyledon specific sequence is characterized by acidic heptameric repeat units containing stretches of glutamic acid, tyrosines and leucines with regular spacing. Other structural motifs, such as a nucleotide binding domain in the subunit and a zinc finger in the subunit, are compared at the amino acid level throughout the RNA polymerase subunits with the enzymes from other organisms in order to identify functionally important conserved regions.The sequence data presented in this paper will appear in the EMBL/Gen Bank/DDBJ Nucleotide Databases under the accession number X17318     

Growth phase-dependent changes in the expression of global regulatory genes and associated metabolic pathways in Escherichia coli

Metabolic regulation in Escherichia coli was studied in terms of the changes in the expression of the global regulatory genes rpoD, rpoS, soxRS, cra, fadR, iclR and arcA at three different growth phases, in batch culture. The expression of rpoS and several rpoS-dependent metabolic pathway genes, such as tktB, talA, fumC, acnA, sucA, acs and sodC, were increased (∼1.5 to 2-fold) as the cells entered the late phase of growth. The changes in the expression of other global regulators and their effects on different metabolic pathway genes were less significant, as compared to rpoS, during the later phases of growth.     

Molecular phylogeny of Old WorldTrifolium (Fabaceae), based on plastid and nuclear markers

Phylogenies of Old WorldTrifolium species were constructed using nucleotide sequence data of the internal transcribed spacers (ITS) of nuclear ribosomal DNA and chloroplast DNA restriction site data from PCR-amplified genes and genic regions (rbcL,trnK, andrpoC1–C2). Biogeography, morphological evolution, and the existing classification forTrifolium were examined. The genusTrifolium is strongly supported as monophyletic, however, only one small section (Chronosemium) is monophyletic, although the data are in conflict regarding its placement. The two largest sections of the genus, Sects.Lotoidea andTrifolium, are not supported as monophyletic, as currently circumscribed. Many members of Sect.Lotoidea are basal within the genus, supporting previously-proposed hypotheses concerning plesiomorphic morphological characters and a Mediterranean-Mideast biogeographic origin of the genus.     

Chlorella chloroplast DNA sequence containing a gene for the large subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase and a part of a possible gene for the β′ subunit of RNA polymerase

The sequence of a 2782 bp fragment of the chloroplast genome of Chlorella ellipsoidea has been determined. The region includes the entire gene (rbcL) for the large subunit (LS) of ribulose-1,5-bisphosphate carboxylase/oxygenase and a sequence (rpoC-like) similar to part of the gene for the subunit of E. coli RNA polymerase which is oriented in same direction as rbcL. The arrangement is rpoC-like — 446 bp — rbcL. The rbcL gene codes for a polypeptide of 475 amino acids whose sequence shows 88% homology with those of tobacco and spinach, 94% homology with that of Chlamydomonas, and 85% homology with that of Anacystis. The putative rbcL promoter sequence has homology with E. coli promoter sequences and its putative terminator sequence is capable of forming a stem-and-loop structure.     

Further studies of the sieve-element plastids of theCaryophyllales includingBarbeuia,Corrigiola, Lyallia,Microtea, Sarcobatus,andTelephium

The sieve-element plastids of 69 species of theCaryophyllales were investigated by transmission electron microscopy. All contained the specific subtype-P3 plastids characterized by a peripheral ring of protein filaments. The presence or absence of an additional central protein crystal and their shape being either polygonal or globular as well as the average sizes of the sieve-element plastids are useful features in the characterization of some families.—Barbeuia contains sieve-element plastids that confirm its placement within thePhytolaccaceae. Lyallia differs fromHectorella by including small starch grains in their sieve-element plastids, which otherwise by their globular crystals negate a closer connection to theCaryophyllaceae. The lack of a central protein crystal in its form-P3fs plastids placesMicrotea best within theChenopodiaceae. Sarcobatus, a so far uncontested member of theChenopodiaceae, contains form-P3cf plastids, i.e., including a central crystal not found elsewhere in this family.Telephium andCorrigiola, shifted back and forth betweenMolluginaceae andCaryophyllaceae, have form-P3cf(s) plastids with a polygonal crystal which favor their placement within theCaryophyllaceae.     

Entwicklungsgeschichte und Ultrastruktur von Pollenkitt und Exine bei nahe verwandten entomophilen und anemophilen Angiospermensippen derAlismataceae,Liliaceae, Juncaceae,Cyperaceae, Poaceae undAraceae

Some closely related members of the monocotyledonous familiesAlismataceae, Liliaceae, Juncaceae, Cyperaceae, Poaceae andAraceae with variable modes of pollination (insect- and wind-pollination) were studied in relation to the ultrastructure of pollenkitt and exine (amount, consistency and distribution of pollenkitt on the surface of pollen grains). The character syndromes of pollen cementing in entomophilous, anemophilous and intermediate (ambophilous or amphiphilous) monocotyledons are the same in principal as in dicotyledons. Comparing present with former results one can summarize: 1) The pollenkitt is always produced in the same manner by the anther tapetum in all angiosperm sub-classes. 2) The variable stickiness of entomophilous and anemophilous pollen always depends on the particular distribution and consistency of the pollenkitt, but not its amount on the pollen surface. 3) The mostly dry and powdery pollen of anemophilous plants always contains a variable amount of inactive pollenkitt in its exine cavities. 4) A step-by step change of the pollen cementing syndrome can be observed from entomophily towards anemophily. 5) From the omnipresence of pollenkitt in all wind-pollinated angiosperms studied one can conclude that the ancestors of anemophilous angiosperms probably have been zoophilous (i.e. entomophilous) throughout.

     

Synopsis of the tribe Bocageeae (Annonaceae), with revisions of Cardiopetalum,Froesiodendron, Trigynaea,Bocagea, and Hornschuchia

The tribal name Bocageeae Endlicher is reestablished and the tribe is circumscribed on the basis of solitary internodal ebracteate pedicels that are articulated at the base, and pollen shed in polyads of eight or more grains. Septate anther locules, large pollen size, and seed appendages are prevalent in the tribe. Intectate pollen with free-standing columellae, rare in Annonaceae, occurs not only in the genusTrigynaea but also in some species ofBocagea andHornschuchia. As defined here, the Bocageeae include seven neotropical genera:Cymbopetalum, Porcelia, Bocagea, Cardiopetalum, Froesiodendron, Hornschuchia, andTrigynaea. The latter five genera are revised and the treatments include ten new species:Cardiopetalum plicatum, Froesiodendron urceocalyx, Hornschuchia lianarum, H. santosii, H. leptandra, Trigynaea cinnamomea, T. lanceipetala, T. triplinervis, T. lagaropoda, andT. axilliflora, all from tropical South America. A new combination,Froesiodendron longicuspe, changes the rank of that taxon from subspecies to species.Cardiopetalum surinamense is removed fromFroesiodendron and reassigned toCardiopetalum on the basis of its connate petals, dehiscent monocarps, and seeds with bilobed arils. A cladogram provides an explicit hypothesis of intergeneric relationships in the tribe. The new combinationOnychopetalum periquino, based onTrigynaea periquino, is made.     

rbcL sequences support exclusion ofRetzia,Desfontainia, andNicodemia from theGentianales

The taxonomic positions ofRetzia, Desfontainia, andNicodemia have been much discussed, and all three genera have been included inLoganiaceae (Gentianales). We have made a cladistic analysis ofrbcL gene sequences to determine the relationships of these taxa toGentianales. Four newrbcL sequences are presented; i.e., ofRetzia, Desfontainia, Diervilla (Caprifoliaceae), andEuthystachys (Stilbaceae). Our results show thatRetzia, Desfontainia, andNicodemia are not closely related toLoganiaceae or theGentianales. Retzia is most closely related toEuthystachys and is better included inStilbaceae. The positions ofDesfontainia andNicodemia are not settled, butDesfontainia shows affinity for theDipsacales s.l. andNicodemia for theLamiales s.l.     

A reconsideration of the genusRabdosiella (Lamiaceae,Nepetoideae, Ocimeae)

The two generaPlectranthus andIsodon are compared and found to be very dissimilar.Isodon ist considered to be misplaced inOcimeae subtribePlectranthinae and apparently is more closely related to subtribeHyptidinae. The disjunct genusRabdosiella is compared to these two genera and regarded to be polyphyletic. The AfricanR. calycina (Benth.)Codd is returned toPlectranthus and calledP. calycinus Benth., while the AsianR. ternifolia (D. Don)Codd is placed inIsodon sect.Pyramidium and calledI. ternifolius (D. Don)Kudo.     

A new fungal genus, Teracosphaeria, with a phialophora-like anamorph (Sordariomycetes, Ascomycota)

The new genus and species Teracosphaeria petroica is described for a perithecial ascomycete and its anamorph occurring on decayed wood collected in New Zealand. The fungus produces immersed, non-stromatic ceratosphaeria-like perithecia in nature, with hyaline, septate ascospores produced in unitunicate, non-amyloid asci. The anamorph produced in vitro is phialophora-like with lightly pigmented phialides terminating in flaring, deep collarettes that are often noticeably brown with conspicuous periclinal thickening. Phylogenetic analysis of LSU rDNA sequence data indicates that this fungus is distinct from morphologically similar fungi classified in the Chaetosphaeriales, the Trichosphaeriales or the Magnaporthaceae. It forms a monophyletic group with recently described, chaetosphaeria-like ascomycetes, such as the pyrenomycete genus Mirannulata, and shows affinity with the anamorphic species Dictyochaeta cylindrospora. The usefulness of describing anamorph genera for morphologically reduced anamorphs, when anamorph characteristics are actually part of the holomorph diagnosis, is discussed. An apparently contradictory example of the so-called Cordana and Pseudobotrytis anamorphs of Porosphaerella spp. is also discussed.     

Species-specific albumin in fossil bones from Orce, Granada, Spain

A skull fragment (VM-0) from Orce, Granada, Spain, dated palaeomagnetically at about 1.6 Myr, is thought by some palaeontologist to be hominid, while others maintain it is equid. If hominid, it would be by far the oldest evidence ofHomo in Europe. Immunological studies on residual albumin in this fossil were carried out independently, and with different immunological methods, at the University of California, San Francisco (radioimmunoassay), and at the University of Granada, Spain (enzyme immunoassay). Other fossils attributed to hominids also studied wereVM1960 from Venta Micena, andCV-1 from Cueva Victoria, Murcia, Spain. Undisputed equid and bovid fossils from the same deposits and dated to a similar period as the Orce skull were also analyzed. Our results showed that species-specific albumin can be detected in 1.6 Myr-old hominid, equid and bovid fossils. The albumin from the Orce skull fragment and fromVM-1960 was immunologically closer to human albumin. These findings support the contention that theVM-0 andVM-1960 are hominid and that members of the genusHomo occupied southern Spain 1.6 Myr ago.     

Novel glycopeptides,containing desmosine and isodesmosine,isolated from porcine aorta

Intima-media of porcine thoracic aorta were digested with pronase, after extraction of the saline-soluble matters and fat. A glycopeptide fraction was precipitated with 90% (vol/vol) ethanol from the 80% ethanol-soluble fraction of the trichloroacetic acid (7%)-soluble fraction of the pronase digest. The glycopeptide fraction was fractionated by affinity chromatography on concanavalin A (Con A)-Sepharose 4B, yielding 4 fractions (FA, FB, FC and FD). The most carbohydrate-rich fraction (FB) was further purified to a homogeneous state. The purified FB (FB-0.1) and all other fractions contained desmosine and isodesmosine. The major sugars in the fractions without or with low affinity for Con A (FA, FB, and FB-0.1) were glucosamine, galactose, mannose and sialic acid, while those in the fractions with high affinity for this lectin (FC and FD) were glucosamine, glucose and mannose. All the fractions contained glycine, aspartic acid (and/or asparagine), serine, proline, threonine, glutamic acid (and/or glutamine) and alanine as the major amino acids, amounting to approximately 80% of the total.     

Triads Synthesized from Deuteroporphyrin IX,Naphthoquinone, and Aromatic Amino Acids

Two triad systems were synthesized from deuteroporphyrin IX by tethering 1,4-naphthoquinone derivative and aromatic amino acids to its propionic groups using the method of mixed anhydrides. Physicochemical characteristics of the triads were studied, and the -electron systems of their chromophores were shown to interact.     

Phylogeny, biogeography, and species boundaries within the Alexandrium minutum group

The geographic range and bloom frequency of the toxic dinoflagellate Alexandrium minutum and other members of the A. minutum group have been increasing over the past few decades. Some of these species are responsible for paralytic shellfish poisoning (PSP) outbreaks throughout the world. The origins of new toxic populations found in previously unaffected areas are typically not known due to a lack of reliable plankton records with sound species identifications and to the lack of a global genetic database. This paper provides the first comprehensive study of minutum-group morphology and phylogeny on a global scale, including 45 isolates from northern Europe, the Mediterranean, Asia, Australia and New Zealand.Neither the morphospecies Alexandrium lusitanicum nor A. angustitabulatum was recoverable morphologically, due to large variation within and among all minutum-group clonal strains in characters previously used to distinguish these species: the length:width of the anterior sulcal plate, shape of the 1′ plate, connection between the 1′ plate and the apical pore complex, and the presence of a ventral pore. DNA sequence data from the D1 to D2 region of the LSU rDNA also fail to recognize these species. Therefore, we recommend that all isolates previously designated as A. lusitanicum or A. angustitabulatum be redesignated as A. minutum. A. tamutum, A. insuetum, and A. andersonii are clearly different from A. minutum on the basis of both genetic and morphological data.A. minutum strains from Europe and Australia are closely related to one another, which may indicate an introduction from Europe to Australia given the long history of PSP in Europe and its recent occurrence in Australia. A minutum from New Zealand and Taiwan form a separate phylogenetic group. Most strains of A. minutum fit into one of these two groups, although there are a few outlying strains that merit further study and may represent new species. The results of this paper have greatly improved our ability to track the spread of A. minutum species and to understand the evolutionary relationships within the A. minutum group by correcting inaccurate taxonomy and providing a global genetic database.     

Identification of b,c, and d cytochromes in the membrane of Vitreoscilla

Cytochromes b, c, d, and o were identified by spectroscopic analysis of respiratory membrane fragments from Vitreoscilla sp., strain C1. Carbon monoxide difference spectra of the reduced membranes had absorption maxima at 416, 534, and 571 nm (ascribed to cytochrome o) and 632 nm (cytochrome d). Derivative spectra of the pyridine hemochromogen spectra of the membranes identified the presence of b- and c-type cytochromes in Vitreoscilla. The cyanide binding curve of the membranes was biphasic with dissociation constants of 2.14 mM and 10.7 mM which were assigned to cytochrome o and cytochrome d, respectively. Membranes bound carbon monoxide with dissociation constant 3.9 M, which was assigned to cytochrome o. Cytochrome c ₅₅₆ and a NADH-p-iodonitrotetrazolium violet reductase component were partially purified from Vitreoscilla membranes.Abbreviations INT p-iodonitrotetrazolium violet - RMF respiratory membrane fragments - K _d dissociation constant - CHAPS 3-[(3-cholamido propyl) dimethylammonio]-1-propanesulfonate - DOC sodium deoxycholate - PAGE polyacrylamide gel electrophoresis - SDS sodium dodecyl sulfate     

Gametogenesis,endosperm development,and embryogeny inMicrocarpaea muscosa (Scrophulariaceae,Gratioleae)

InMicrocarpaea muscosa gametogenesis, embryo sac development (Polygonum type), endosperm development (cellular), embryogeny (Onagrad type), and seed structure are described. The different modes of endosperm development withinGratioleae are compared.