克雷伯氏菌属(Klebsiella)细菌比较基因组学分析揭示多复制子抗性质粒介导广泛的抗性基因传播

[目的] 研究克雷伯氏菌与多复制子抗性质粒间的关系,分析细菌携带多复制子质粒对抗生素环境的响应机制。[方法] 以2018-2020年分离的56株不同来源克雷伯氏菌（Klebsiella sp.）分离株为研究对象,利用微量肉汤稀释法评估其多重耐药表型,对分离菌株进行全基因组测序（WGS）,通过细菌全基因组关联分析（BGWAS）技术和比较基因组学方法深入解析多复制子抗性质粒形成的机制。[结果] 耐药表型分析发现野生动物来源的菌株具有更广的耐药谱系,总体Klebsiella sp.对氨苄西林表现出很高的耐药率（80.36%）,尤其是马来穿山甲来源菌株对头孢类抗生素高度耐受,同时对氯霉素、左氧氟沙星和复方新诺明等药物耐受,基因组分析发现这些菌株携带了抗性质粒和更多的抗生素抗性基因。进一步对69个质粒序列分析,发现有28个质粒为多复制子质粒,主要携带bla_CTX-M-15、bla_CTX-M-14、bla_CTX-M-55、bla_OXA-1和bla_TEM-1等β-内酰胺酶基因。细菌携带质粒类型分析认为Klebsiella pneumoniae可能是多复制子质粒的重要宿主,质粒骨架与结构分析发现多复制子质粒多由2个或2个以上单个质粒融合而成,携带此类质粒的菌株不仅获得了更广的耐药表型,而且在全球传播扩散分布逐年增加,因此产生对抗生素环境更强的适应性。[结论] 多重耐药性细菌呈现的表型与携带的多复制子质粒有关,相比较下多复制子质粒比非多复制子质粒有更强的抗性基因携带能力,或许是细菌在强大的抗生素压力下产生的重要响应机制。本研究对于未来探索细菌抗性基因的传播扩散机制具有重要意义。     

The native subspecies of Juncus effusus (Juncaceae) in western North America

Juncus effusus s. str. has two native representatives on the coast of western North America. A new combination is provided forJuncus effusus subsp.pacificus, ranging from California to British Columbia.Juncus effusus subsp.austrocalifornicus is newly described from southernmost California, Arizona, and Baja California, México. It differs from subsp.pacificus by its pale sheaths, with less thickened and less overlapping margins, and its disjunct distribution. A key is provided for the subspecies ofJ. effusus in North America.     

A survey of the genus Festuca L. (Poaceae) in Italy. Festuca gamisansii Kerguélen subsp. aethaliae,subsp. nova

ABSTRACT

Festuca gamisansii subsp. aethaliae, endemic to the Isle of Elba (Tuscan Archipelago) is described. The morphological, anatomical, karyological, ecological and chorological characteristics of the species are given. The main characteristics distinguishing F. gamisansii subsp. aethaliae from F. gamisansii subsp. gamisansii occurring in Corsica are also summarized.     

浅紫灰链霉菌CQ01819及其次级代谢产物的定向发掘

【目的】采用特征次级代谢产物生物合成的保守功能基因探针,定向分离土壤中产生特征次级代谢产物的菌种资源,借助基因转录分析为导向的培养基优化方法,获得目标次生代谢产物。【方法】首先,根据5种特征次级代谢产物保守的合成功能基因设计简并引物,定向从土壤样品中筛选、分离并纯化菌株。然后,以RT-qPCR为指导开展目标产物的发酵培养基优化;最后,对菌株进行发酵,利用多种色谱技术分离纯化目标天然产物,并结合高分辨质谱与核磁共振等技术对所获得的化合物进行结构鉴定。【结果】从土壤中筛选得到了一株AHBA合酶基因和环氧化酶基因均为阳性的链霉菌菌株(编号为CQ01819),根据转录分析优化发酵培养基,最终从该菌株分离纯化得到了含有AHBA结构单元的丝裂霉素C、聚醚类抗生素莫能霉素A和缬吲霉素。【结论】本研究通过菌株的定向分离纯化,筛选得到了产生预期抗生素的浅紫灰链霉菌菌株CQ01819;基于RT-qPCR指导的发酵培养基优化,确定了菌株的发酵条件;获得发酵粗提物后,采用多种色谱整合技术和光谱分析策略,快速分离并鉴定了目标产物。该研究为目标菌种资源的定向筛选、菌株的发酵条件的快速优化和化合物的定向分离提供了较...     

Structure-Activity Studies on Monoamine Oxidase Inhibitors by Calorimetric and Quantum Mechanical Calculations

Abstract

Structure-activity relationship studies were carried out on a new series of hydrazino-thiosemicarbazide derivatives, which inhibit monoamino oxidase (MAO). Fifty-five compounds were synthesized and tested “in vitro” for their inhibitory effects on rat liver mitochondrial MAO. The most efficient MAO inhibitors were the benzylidene derivatives (R-CH=N1-N2H-C=N4R1-SR2) where R is the piperonyl radical and ethyl or isopropyl substituents are in R1 position. Correlation of MAO activity with hydrophobic, electronic and steric properties of tested compounds, evaluated by means of Quantum Mechanical calculations and calorimetric analysis (DSC) suggest that electronic and steric parameters give a better fit than hydrophobicity with the biological activity.     

Design,synthesis of novel pyranotriazolopyrimidines and evaluation of their anti-soybean lipoxygenase,anti-xanthine oxidase,and cytotoxic activities

The synthesis of 14-(aryl)-14H-naphto[2,1-b]pyrano[3,2-e][1,2,4]triazolo[1,5-c]pyrimidine-2-yl) acetamidoximes 2a–e has been accomplished by reaction of 2-acetonitrile derivatives 1a–e with hydroxylamine. Cyclocondensation reaction of precursors 2a–e with some elctrophilic species such as ethylorthoformate, acetic anhydride, and methyl-acetoacetate provided the new oxadiazole derivatives 3a–e, 4a–e, and 5a–e, respectively. On the other hand, the reaction of precursors 2a–e with 2-chloropropanoyl chloride afforded the new acetimidamides 6a–e which evolve under reflux of toluene to the new oxadiazoles 7a–e. The synthetic compounds were screened for their anti-xanthine oxidase, anti-soybean lipoxygenase, and cytotoxic activities. Moderate to weak xanthine oxidase and soybean lipoxygenase inhibitions were obtained but significant cytotoxic activities were noted. The most cytotoxic activities were recorded mainly (i) 5a was the most active (IC_50?=_?4.0?μM) and selective against MCF-7 and (ii) 2a was cytotoxic against the four cell lines with selectivity for MCF-7 and OVCAR-3 (IC_50?=_?17 and 12?μM, respectively) while 2e is highly selective against OVCAR-3 (IC_50?=_?10?μM).     

Note critiche sulla flora d'Italia. II. Il gruppo di Minuartia verna

Abstract

Critical notes on the flora of Italy. II. The Minuartia verna group. The following paper contains a re-examination of the Minuartia verna–complex in Italy based on a revision of herbarium-specimens and observations in nature. The author proposes to divide this polymorphic species into 4 subspecies. Two of them (subsp. verna and subsp. collina) are largely corresponding to the opinion of earlier authors. The subsp. attica is also represented in the flora of Southern Italy, but Italian populations are slightly different from those of Greece (which must be regarded as typical). The subsp. grandiflora living in a small area in Sicily, which was incompletely known, is described and its relations to the other subspecies are discussed.     

THE GEOGRAPHICAL DISTRIBUTION OF B-CHROMOSOMES OF XANTHISMA TEXANUM DC. (ASTERACEAE). I. SURVEY OF THE RANGE

The distribution of B-chromosomes throughout the range of Xanthisma texanum has been determined from approximately 750 individuals from 80 wild populations in Texas, Oklahoma, and New Mexico. Counts of meiotic chromosome figures of n = 4_II + 1 B_I and n = 5_II (4_II + IB_II) are reported for the first time for subsp. texanum var. texanum and for subsp. texanum var. orientalis. The wide occurrence of B-chromosomes in natural populations of subsp. drummondii is also documented. B-chromosomes are present in 22 (27.2%) randomly distributed populations whose characteristics are in no way distinct from populations without B-chromosomes. In populations with B-chromosomes present, 19.8% of the plants contained one or two B-chromosomes, but in total only 5.8 % of all plants contained one or two B-chromosomes. All populations in one isolated population group of subsp. drummondii lack B-chromosomes.     

Segnalazione di piante nuove,inedite, o notevoli per la regione marchigiana. III

Abstract

A report about new or interesting plants in the Marche region. III. — The authors report about the occurrence in the Marche of 61 entities of specific or infraspecific rank, 22 of which are new or unpublished concerning the region, while the remaining 39 have been previously very little or imprecisely mentioned; as to the latter new localities and stations are shown or general information on the distribution over the Marche district has been supplied. Out of the first 22 entities, 18 (Fumaria muralis subsp. muralis, Cardamine flexuosa, Lunaria annua subsp. pachyrhiza, Colutea arborescens subsp. gallica, Astragalus monspessulanas subsp. illyricus, Cornus sanguinea subsp. australis, Galium verrucosum, Hedypnois rhagadioloides subsp. tubaeformis, Andryala tenuifolia, Sonchus asper subsp. glaucescens, Rheichardia picroides var. maritima, Podospermum canum, Scorzonera austriaca, S. hirsuta, Bidens frondosa, Guizotia abyssinica, Ambrosia elatior, A. trifida) are, as far as the authors know, quite new or unpublished concerning the Marche, while 4 (Rumex angiocarpus, Trifolium repens subsp. prostratum, Epilobium lanceolatum, Picris hieracioides subsp. spinulosa) are generally reported in Middle Italy or over the whole Italian territory by the general Floras, but till now have not been mentioned in works dealing with regional flora.     

内蒙古呼伦贝尔地区传统发酵乳中乳酸菌的多样性分析

【目的】对内蒙古呼伦贝尔地区传统发酵乳制品中乳酸菌资源的生物多样性进行研究。【方法】采用纯培养和16S rRNA基因序列分析法对内蒙古呼伦贝尔地区传统发酵乳中的乳酸菌进行多样性分析。【结果】从8份传统发酵乳制品(6份酸牛奶和2份酸马奶)样品中分离到24株乳酸菌,通过16S rRNA基因序列分析和系统进化关系分析将24株乳酸菌鉴定为2株Lactobacillus kefiranofaciens、2株Lactobacillus kefiri、5株Lactobacillus paracasei、3株Lactobacillus plantarum、1株Lactobacillus rhamnosus、6株Lactococcus lactis subsp.lactis、2株Leuconostoc mesenteroides subsp.dextranicum、2株Streptococcus thermophilus和1株Enterococcus faecium。【结论】Lactococcus lactis subsp.lactis为内蒙古呼伦贝尔地区传统发酵乳制品的优势菌种,占总分离株的25%,其次为Lactobacillus paracasei,占总分离株的20.83%。     

Production of NO and N2O by the Heterotrophic Nitrifier Alcaligenes faecalis parafaecalis under Varying Conditions of Oxygen Saturation

Production of NO and N ₂ O by the heterotrophic nitrifier Alcaligenes faecalis subsp. parafaecalis was studied during growth in batch and continuous culture on peptone-meat extract medium. Depending on oxygen saturation level, medium redox status and amount of substrate supplied, the microorganisms produced 0.002–0.25 mg NO-N h^{- 1} (g protein)^{- 1} and 0.16–2.4 mg N ₂ O-N h^{- 1} (g protein)^{- 1} . Maximum rates of nitrogen oxides production were observed during peak events initiated by sudden changes of oxygen supply in the medium and were due to combined nitrification/denitrification taking place simultaneously within the cells. Based on model simulations of enzymatic kinetics of denitrification, possible mechanisms of increased nitrogen oxides production during periods of changes in oxygen supply are suggested.     

Effect of cell membrane-active antimicrobial agents on membrane potential and viability of mycoplasmas

The membrane potential ofMycoplasma mycoides subsp.capri has been determined to beE _M=−48 mV±10%, inside negative. In this study we investigated the influence of cell membrane-active antimicrobial agents, viz., valinomycin, gramicidin, polymyxin, and clotrimazole, on membrane potential and viability ofM. mycoides subsp.capri. Valinomycin, an ionophore with extreme potassium selectivity, induced a membrane hyperpolarization,E _M=−110 mV. Valinomycin was not cidal, but static to mycoplasmas. Obviously the potassium drain induced by valinomycin can be compensated for by the organisms. Gramicidin is an antibiotic forming cation conduction channels across membranes. It induced a rapid depolarization,E _M=+23 mV, of mycoplasma membranes. At low concentrations, gramicidin had a static effect, whereas at high concentrations it was cidal to mycoplasmas. The rapid permeation of cations through the stationary ion channels formed by gramicidin obviously exerts an inhibitory or even lethal effect on mycoplasma metabolism and growth. Polymyxin B induced a depolarization,E _M=−35 mV, of mycoplasma membranes only when the organisms had been pretreated and hyperpolarized with valinomycin. After treatment with both valinomycin and polymyxin B, a slight inhibition of mycoplasma growth was observed. Clotrimazole, a synthetic imidazole antimycotic, hyperpolarized mycoplasma membranes (E _M=−80 mV). At high concentrations clotrimazole was cidal, whereas at low concentrations it was static to mycoplasmas.     

Parallel Evolution Under Domestication and Phenotypic Differentiation of the Cultivated Subspecies of <Emphasis Type="Italic">Cucurbita pepo</Emphasis> (Cucurbitaceae)

Parallel Evolution under Domestication and Phenotypic Differentiation of the Cultivated Subspecies of Cucurbita pepo (Cucurbitaceae). Cucurbita pepo (pumpkin, squash, gourd, Cucurbitaceae) is an ancient North American domesticate of considerable economic importance. Based on molecular genetic polymorphisms, two cultivated lineages of this species, each consisting of very many edible–fruited cultigens, have been recognized, C. pepo subsp. pepo and C. pepo subsp. texana. However, the phenotypic commonalities and differences between these two subspecies have not as yet been systematically collected and organized. Among the evolutionary developments common to the two subspecies are the increased size of the plant parts, less plant branching, and premature loss of chlorophyll in the exocarp of the fruits. In both subspecies, bush growth habit, conferred by allele Bu, is common to the cultigens grown for consumption of the immature fruits, as is the deviation from the 1:1 ratio of fruit length to fruit width. A major characteristic differentiating between the edible–fruited cultigens of the respective subspecies are the longitudinal protrusions, in subsp. pepo, versus depressions, in subsp. texana, of the fruit surface corresponding with the subsurface primary carpellary vein tracts. Subsp. pepo also has larger fruits and larger and longer seeds. In addition, some alleles affecting stem color, leaf mottling, multiple flower bud production, and fruit characteristics are frequently occurring to nearly fixed in one subspecies but are rare to less common in the other.     

Magnolia (Magnoliaceae) in Mexico and Central America: a synopsis

As a result of a recent alpha-taxonomic study of the genusMagnolia s. str. in Mexico and Central America, four new species and four new subspecies ofMagnolia sect.Theorhodon are proposed, includingM. iltisiana, M. panamensis, M. tamaulipana, M. sororum subsp.lutea, andM. pacifica, consisting of three subspecies:pacifica, pugana, andtarahumara. Additionally, a new combination,M. guatemalensis subsp.hondurensis, is made, and a new section,M. sect.Splendentes, is erected for the Caribbean species. The results have implications for understandingMagnolia biogeography, andM. tamaulipana has potential for horticulture in temperate zones. Keys to genera, sections, and species, and maps of species distributions are presented.Magnolia grandiflora L. andM. schiedeana Schltdl. are lectotypified. Como resultado de un estudio alfa-taxonómico reciente del géneroMagnolia s.str. en México y Centro América, se proponen cuatro especies nuevas y cuatro subespecies nuevas deMagnolia sect.Theorhodon, incluyendoM. iltisiana, M. panamensis, M. tamaulipana, M. sororum subsp.lutea y.M. pacifica, la última consiste de tres subespecies:pacifica, pugana ytarahumara. Adicionalmente, se hace la combinaciónM. guatemalensis subsp.hondurensis y se erigeM. sect.Splendentes para las especies del Caribe. Los hallazgos tienen implicaciones en el entendimiento de la biogeografia deMagnolia y.M. tamaulipana tiene potencial en la horticultura de zonas templadas. Se presentan claves para géneros, secciones y especies, y mapas de distribución para las especies.Magnolia grandiflora L. yM. schiedeana Schltdl. son lectotipificadas.     

Segnalazione di Piante Nuove,Inedite o Notevoli per la Regione Marchigiana. I

Abstract

A report about new or interesting plants in the Marche region. I. — The author points out or in some cases confirms the occurrence in this region of 107 species or subspecies. The report is based on the collections and observations made by the author himself and his collaborators during the last years. 31 of such entities (Asplenium trichomanes subsp. quadrivalens, Amaranthus blitoides, Cerastium brachypetalum subsp. roeseri, C. silvaticum, Silene alba subsp. divaricata, Ceratocephalus falcatus, Malcolmia orsiniana, Arabis collina, Fibigia clypeata, Neslia paniculata subsp. thracica, Hymenolobus pauciflorus, Biscutella didyma, Hirschfeldia incana, Rapistrum rugosum subsp. orientale, R. rugosum subsp. linnaeanum, Sempervivum italicum, Lembotropis nigricans, Chamaecytisus polytrichus, Ononis mitissima, Melilotus neapolitana, Trifolium hybridum subsp. hybridum, T. phleoides, Lotus edulis, Oxalis europaea, O. articulata, Euphorbia prostrata, Rhamnus alpinus subsp. fallax, Fumana arabica, Oenonthe lachenalii, Pimpinella tragium subsp. lithophila, Ferula communis subsp. glauca) are quite new or never dealt with for the Marche region, sometimes even for the whole central section of the Italian Peninsula; other 18 (Moehringia pentandra, Minuartia mediterranea, Stellaria nemorum subsp. glochidisperma, Cerastium semidecandrum subsp. semidecandrum, Sagina maritima, Spergularia marina, S. bocconii, Petrorhagia velutina, Adonis flammea, Sisymbrium orientale, Rorippa amphibia, Cardamine resedifolia, Capsella rubella, Potentilla detomasii, Vicia lathyroides, Trigonella monspeliaca, Hippocrepis ciliata, Geranium purpureum) are briefly reported in Middle Italy by the Italian general Floras, but till now have not been mentioned in works dealing with regional floristic. The remaining 58 entities are already known in the Marche, but scarcely reported, or their location is either not clearly defined or insufficiently described. For these new localities and stations are given, or general information on their distribution throughout the region is provided.     

Genetic diversity in the endangered Sicilian endemic Brassica rupestris: Proposals for a conservation strategy

Abstract

Brassica rupestris Raf. is a chasmophyte species that includes two subspecies, both endemic to Central-Western Sicily (Italy). Inter-Simple Sequence Repeat (ISSR) markers were used to detect genetic diversity within and among eight populations representative of the species' distribution range. High levels of genetic diversity were revealed both at the population (PPB = 53.88%, H _S = 0.212, Sh = 0.309) and at the species level (PPB = 96.55%, H _T = 0.307, Sh = 0.464). The correlation between genetic and geographical distances was negative (Mantel test, r = ?0.06, P < 0.95). The two subspecies of B. rupestris, subsp. rupestris and subsp. hispida, showed remarkable genetic similarity and molecular data did not unequivocally support their distinctness. The pattern of genetic variation revealed by our study bears important consequences for conservation management: It is desirable to preserve B. rupestris populations in situ with a “dynamic” strategy, while, ex situ conservation programmes might be improved to safeguard maximum genetic diversity.     

Seed germination in the Sicilian subspecies of Dianthus rupicola Biv. (Caryophyllaceae)

Abstract

Five populations belonging to three subspecies of Dianthus rupicola Biv. (D. rupicola subsp. rupicola, D. rupicola subsp. aeolicus, and D. rupicola subsp. lopadusanus) and growing in different geographical areas of Sicily were tested for seed germination at various temperatures. All populations showed high germination rates with an optimum temperature between 15°C and 25°C. Efficient seed germination might contribute significantly to the preservation of these subspecies which are currently exposed to several environmental threats.     

Kinetic studies on the reactions of Fusarium galactose oxidase with five different substrates in the presence of dioxygen

 Reactions (25  °C) of galactose oxidase, GOase_ox from Fusarium NRRL 2903 with five different primary-alcohol-containing substrates RCH₂OH:- D-galactose (I) and 2-deoxy-d-galactose (II) (monosaccharides); methyl-β-d-galactopyranoside (III) (glycoside);d-raffinose (IV) (trisaccharide); and dihydroxyacetone (V) have been studied in the presence of O₂. The GOase_ox state has a tyrosyl radical coordinated at a square-pyramidal Cu^II active site, and is a two-equivalent oxidant. Reactant concentrations were [GOase_ox] (0.8–10 μM), RCH₂OH (1.0–6.0 mM), and O₂ (0.14–0.29 mM), with I=0.100 M (NaCl). The reactions, monitored at 450 nm by stopped-flow spectrophotometry, terminated with depletion of the O₂. Each trace was fitted to the competing reactions GOase_ox+RCH₂ OH → GOase_redH₂+RCHO (k ₁), and GOase_redH₂+O₂→ GOase_ox+H₂O₂ (k ₂), with GOase_redH₂ written as the doubly protonated two-electron-reduced Cu^I product. It was necessary to avoid auto-redox interconversion of GOase_ox and GOase_semi . Information obtained at pH 7.5 indicates a 5 : 95 (ox : semi) "native" mix equilibration complete in ∼3 h. At pH >7.5, rate constants 10^–4 k ₁ / M^–1 s^–1 for the reactions of GOase_ox with (I) (1.19), (II) (1.07), (III) (1.29), (IV) (1.81), (V) (2.94) were determined. On decreasing the pH to 5.5, k ₁ values decreased by factors of up to a half, and acid dissociation pK _as in the range 6.6–6.9 were obtained. UV-Vis spectrophotometric studies on GOase_ox gave an independently determined pK _a of 6.7. No corresponding reactions of the Tyr495Phe variant were observed, and there are no similar UV-Vis absorbance changes for this variant. The pK _a is therefore assigned to protonation of Tyr-495 which is a ligand to the Cu. The rate constant k ₂ (1.01×10⁷ M^–1 s^–1) is independent of pH in the range 5.5–9.0 investigated, suggesting that H⁺ (or H-atoms) for the O₂ → H₂O₂ change are provided by the active site of GOase_red . The Cu^I of GOase_red is less extensively complexed, and a coordination number of three is likely. Received: 4 February 1997 / Accepted: 16 May 1997