The production of two Th2 cytokines, interleukin-4 and interleukin-10, is controlled independently by locus Cypr1 and by loci Cypr2 and Cypr3, respectively

 The strains BALB/cHeA (BALB/c) and STS/A (STS) differ in production of IL-4 and IL-10, two Th2 cytokines, after stimulation of spleen cells with Concanavalin A, STS being a low and BALB/c a high producer. We analyzed the genetic basis of this strain difference using the recombinant congenic (RC) strains of the BALB/c-c-STS/Dem (CcS/Dem) series. This series comprises 20 homozygous strains. Each CcS/Dem strain contains a different, random set of approximately 12.5% genes of the "donor" strain STS and approximately 87.5% of the "background" strain BALB/c. We selected for further analysis the RC strain production intermediate between BALB/c and STS. In (CcS-20×BALB/c)F₂ hybrids we found that different loci control expression of IL-4 and IL-10. Cypr1 (cytokine production 1) on chromosome 16 near D16Mit15 controls IL-4 production, whereas the production of IL-10 is influenced by loci Cypr2 near D1Mit14 and D1Mit227 on chromosome 1 and Cypr3 marked by D5Mit20 on chromosome 5. In addition, the relationship between the level of these two cytokines depends on the genotype of the F₂ hybrids at a locus cora1 (correlation 1) on chromosome 5. This differential genetic regulation may be relevant for the understanding of biological effects of T-helper cells in mice of different genotypes. Received: 2 March 1998 / Revised: 8 June 1998     

A genetic assay for transversion mutations in bacteriophage T4

Summary Seventy-two mutants deficient in formate-nitrate reductase activity were selected in Escherichia coli strain PK 27, by two different procedures. Forty-five strains were selected on the basis of chlorate resistance and 27 strains were selected by their inability to reduce nitrate with formate as an electron donor. Genetic analysis of these strains showed that the two techniques yield distinctly different distributions of mutants among the various controlling genetic loci. Chlorate resistance appears to select for severe alterations in the nitrate reductase system; 98% of these mutants fell into the pleiotropic chl A, B, D and E classes and are deficient in all the activities of the formate-hydrogenlyase pathway as well as formate-nitrate reductase pathway. In contrast, 48% of the mutants selected for their inability to reduce nitrate with formate as the electron donor were of the chl C class and two new classes were identified among mutants selected by this procedure. Chl F mutants are linked to tryptophan and the chl C locus. Chl G mutants map at zero minutes on the E. coli genetic map.     

The use of a phospholipase A-less Escherichia coli mutant to establish the action of granulocyte phospholipase A on bacterial phospholipids during killing by a highly purified granulocyte fraction

Phospholipase A₂ present in a highly purified, potently bactericidal, fraction from rabbit granulocytes produces net bacterial phospholipid degradation during killing of a phospholipase A-less strain of Escherichia coli. In the wild-type parent strain phospholipid breakdown is caused not only by the action of phospholipase A₂ but also by phospholipase A₁, indicating activation of the most prominent phospholipase of E. coli. This activation occurs as soon as the bacteria are exposed to the granulocyte fraction. Phospholipid breakdown by both phospholipases A is dose dependent but reaches a plateau after 30–60 min and at higher concentrations of the fraction.Phospholipid degradation is accompanied in both strains by an increase in permeability to actinomycin D that is also dose dependent. Even though net hydrolysis of phospholipids is greater in the parent strain than in the mutant, the increase in permeability is the same in the two strains.The addition of 0.04 M Mg²⁺, after the effects on phospholipids and permeability have become manifest, initiates in both strains the restoration of insensitivity to actinomycin D, the net resynthesis of phospholipids, and the disappearance of monoacylphosphatides and the partial disappearance of free fatty acids that had accumulated. Loss of ability to multiply is not reversed by Mg²⁺ in either strain. Less than 5 μg of granulocyte fraction causes loss of viability of from 90 to 99% of $\text{1 × 10}{}^8$ microorganisms of both strains. However, at lower concentrations the parent strain is considerably more sensitive to the bactericidal effect of the granulocyte fraction than the mutant strain.     

Sporulation in saprotrophic and clinical strains of <Emphasis Type="Italic">Aspergillus sydowii</Emphasis> (Bain. &; Sart.) Thom &; Church under various environmental conditions

Molecular genetic characteristics and capacity for sporulation under different levels of temperature and humidity were compared for three saprotrophic and four clinical strains of A. sydowii. Analysis of the ITS and D1/D2 loci of these A. sydowii strains revealed two clades, each including both the clinical and saprotrophic strains. The differences in sporulation in the saprotrophic and clinical strains of the potentially pathogenic microscopic fungus A. sydowii under different environmental conditions were demonstrated. In the clinical A. sydowii strains, the level of spore formation was generally higher, especially at humidity levels of 0.90 and 0.95 a_w and 20–25°C. The level of spore formation for the clinical strains inoculated into sterile soil was several times higher than for the saprotrophic ones. On the contrary, nonsterile soils (sod-podzolic and urban soils) exhibited a fungistatic effect against A. sydowii populations.     

Decreased membrane integrity in castor bean mitochondria by hydrogen peroxide. Evidence for the involvement of phospholipase D.

Purified mitochondria from germinating castor bean (Ricinus communis L.) endosperm was treated with hydrogen peroxide (H₂O₂), active oxygen form, in order to investigate the extent of membrane degradation. Incubation of mitochondria with micromolar concentrations (50–200 μM) of H₂O₂ resulted in a concentration-dependent loss of membrane proteins. During this process extensive loss of lipid-phosphate content was also observed in mitochondrial membranes. When L-3-phosphatidyl[2-¹⁴C]ethanolamine was added to the mitochondrial membranes as an exogenous substrate, the level of radioactivity in the water-soluble fraction was markedly enhanced with increasing concentration of H₂O₂. Analysis of the water-soluble products formed during the metabolism of ethanolamine-labelled phosphatidylethanolamine by mitochondrial membranes from castor bean indicates that this loss of lipid-phosphate is attributable to action of phospholipase D. Direct measurement of mitochondrial phospholipase D indicated that the activity of enzyme was remarkably stimulated by calcium ion or sodium dodecylsulfate (SDS). The optimum concentrations for enzyme stimulation were 25 and 0.5 mM for calcium ion and SDS in the reaction mixture, respectively. The substrate specificity of phospholipase D was determined by comparing various classes of exogenous phospholipids, added in the form of sonicated vesicles, as substrates. The phospholipase D exhibited preference for phosphatidylethanolamine. Taken together, our results suggest that increase of mitochondrial phospholipase D activity may be a key event leading to accelerated membrane deterioration following active oxygen attack.     

Propionibacterium species diversity in anaerobic digesters

Phenotypic characteristics of 60 strains ofPropionibacterium isolated from anaerobic hybrid digesters treating landfill leachate and a baker's yeast factory effluent were analysed using numerical taxonomy. With the use of the S_SM similarity coefficient, 92% of the anaerobic digester strains were grouped in eight major clusters. The isolates were identified by relating them to specific type strains and comparison of phenotypic characteristics. These clusters were equated with the classical speciesP. acidipropionici, P. freudenreichii, P. jensenii andP. thoenii using the current classification system. Some of the digester isolates were identified to specifies level using the current identification system, but based on overall similarity they were clustered among members of another species. Furthermore, the data indicated that there was low similarity between the digester isolates and the type strains ofP. jensenii andP. thoeni. A hypothesis is presented as to the role of these propionic acid-producing bacteria during the granulation process found in anaerobic digesters.     

耐镉促生菌株的分离鉴定及对大豆的促生效应

【背景】华南地区镉（Cd）污染严重,与有益微生物共生能够使作物通过直接或间接的机制解除镉毒,提高抗逆性,进而促进生长。耐镉促生菌剂具有广泛的应用前景。【目的】从华南地区受镉污染植株的根内和根际筛选出耐镉且能促进大豆生长的促生菌,以丰富促进田间大豆生产的优异菌种资源。【方法】采用平板划线法从植株的根内或根际分离菌株,通过生理生化特性和16S rRNA基因序列分析对分离菌株进行初步研究,利用盆栽试验探究镉胁迫下菌株对大豆生长的影响,通过测定丙二醛含量和总抗氧化能力探究菌株的耐镉机制。【结果】分离获得4株菌D1、D2、D3和D4,促生特性试验证明4株菌均具有溶磷、产吲哚乙酸和铁载体的能力。经16S rRNA基因序列分析鉴定,D1、D2、D3和D4菌株分别属于不动杆菌属、微小杆菌属、类芽孢杆菌属和普罗威登斯菌属。用这4株菌进行不同镉处理的大豆（巴西10号）盆栽试验,结果表明,4株菌均具有耐镉和促进大豆生长的作用。不添加镉的条件下,大豆接种D4菌株的地上部干重、根部干重和株高分别增加了28%、35%和31%;在添加20mg/kg-CdCl₂·5/2H₂...     

Exotoxic activities ofCorynebacterium pseudotuberculosis

Several methods were used to investigate the relationships between staphylococcal beta-hemolysin inhibitory (BHI) activity, phospholipase D, dermonecrosis, and lethality, which have all been used as indicators of exotoxin in culture filtrate ofCorynebacterium pseudotuberculosis. Culture filtrate was subjected to treatment with formalin, heat, and fractionation by (NH₄)₂SO₄ precipitation, ultrafiltration, and Sephadex G-100 chromatography. Formalin treatment of culture filtrate resulted in loss of phospholipase D and dermonecrotic activities, but no decrease in BHI activity. Culture filtrate and formalin-treated culture filtrate blocked complement-mediated immune hemolysis. Heat treatment of culture filtrate destroyed phospholipase-D activity and reduced BHI activity. Phospholipase-D and dermonecrotic activities eluted from Sephadex G-100 as two distinct adjacent peaks preceding the main protein peak. Maximum lethal activity did not correspond to phospholipase D, but was closely associated with dermonecrotic and maximum BHI activity. Dermonecrotic and BHI, but not phospholipase-D activities were detected in a 1000-dalton filtrate of culture filtrate.     

Intrinsic differences in the perturbing ability of alkanols in bilayer: Action of phospholipase A2 on the alkanol-modified phospholipid bilayer

Summary The kinetic parameters for the steady-state rate of hydrolysis of egg phosphatidylcholine in multilamellar vesicles by bee venom phospholipase A₂ are measured in the presence of 27 alkanols and several organic solvents. In general, small nonpolar solutes like enflurane, tetrahydrofuran, benzene, chloroform and diethylether do not promote the hydrolysis of multilamellar vesicles. The rate of hydrolysis shows a biphasic dependence upon the alkanol concentration for all higher (C₅–C₉) alcohols examined, i.e., an optimal rate of hydrolysis is observed at a characteristic concentration for each alcohol. The alkanol to lipid mole ratio (D/L ratio) in the bilayer at the peak activating concentration of an alkanol was computed from its bilayer/water partition coefficient. The branched chain alcohols induce peak activation of hydrolysis at lowerD/L ratios in the bilayer than the corresponding straight chain analogs. Similarly, the longer chainn-alkanols at peak activating concentration have a lowerD/L ratio than the corresponding lower alcohols. Both theK _m andV _m for phosphatidylcholine increase as a function of the chain length of the activating alcohol. These kinetic parameters also depend upon the position of the substituents on the activating alcohols. Both theD/L ratio andV _m for an alcohol are found to change with the cross-sectional area of the activating alcohol across its long axis: alcohols with a more asymmetric cross-section exhibit higherV _m and a lowerD/L ratio. Such correlations ofV _m andD/L ratio with the molecular parameters of the alkanols are interpreted to suggest that the accessibility of the substrate molecule in the bilayer to the phospholipase is modulated by the free space introduced by the alkanols in the bilayer.Effect of tetradecane derivatives and A₂C (a membrane fluidizing agent) on the phase transition characteristics of DPPC bilayers, and their susceptibility to phospholipase A₂ from bee venom and pig pancreas is also reported. These solutes cause a broadening of the transition profile and reduce the size of the cooperative unit and the enthalpy of transition. These effects depend upon the mole fraction of a solute in the bilayer; however, equal concentrations of these solutes do not induce equal response. Susceptibility of the modified bilayers to phospholipase A₂ depends not only upon the structure of the solute but also upon the source of the enzyme. The data show that the activity of the membrane-bound enzyme is modulated to different extents by different solutes, and the bilayer perturbing ability of these solutes may be related to the asymmetry of their cross-sectional area and to the free space introduced by the alkanols in a bilayer.     

Phospholipase activity in Cryptococcus neoformans

Phospholipases have only been detected in a few fungi and yeasts, in particular in Candida albicans. Secreted phospholipases are considered by some researchers to be a potential factor of virulence and pathogenicity in C. albicans. Twenty-three Cryptococcus neoformans strains were tested in order to observe phospholipase production. Twenty-two of the 23 strains tested were able to produce phospholipases, and the ratio diameter of the colony to total diameter of the colony plus zone of precipitation (Pz) ranged between 0.271 and 0.949. C. neoformans, just like C. albicans, can be divided on the basis of the Pz into different strains according to their virulence and pathogenicity. There also appeared to be a correlation between the phospholipase production and the size of the capsule in the strains isolated from AIDS patients. For this reason, further studies on C. neoformans phospholipase activity would be useful in evaluating the virulence of different strains.     

Synthesis of 5-Alkyl-5-methyl-2-hydroxy-2-cyclopentenones

A hydrogen bacterium strain, N34, and its oxygen-resistant segregant strain, Y38, were subjected to a taxonomical study. Since both strains were capable of N₂-fixation, N₂-fixing facultative hydrogen autotrophs listed in “Bergey’s Manual of Systematic Bacteriology” were used for comparison. Both strains produced a water-insoluble carotenoid pigment, zeaxanthin dirhamnoside, indicating that both should be classified into the genus Xanthobacter. Then, the differential characteristics of the two species of the genus Xanthobacter, X. autotrophicus and X. flavus, were investigated as to both strains. The vitamin requirement, the sensitivity to oxygen under autotrophic conditions, the inducibility of hydrogenase, the substrate range of carbohydrates and N₂-fixing growth characteristics of both strains were almost completely opposite to those of X. flavus. Moreover, both strains coincided exactly with X. autotrophicus in morphological and other physiological characteristics. From these results both strains were identified as Xanthobacter autotrophicus.     

SELECTION OF RESISTANCE STRAINS IN LIPOSCELIS BOSTRYCHOPHILA BADONNEL TO CO2-ENRICHED ATMOSPHERES*

Abstract Adults of the psocid Liposcelis bostrychophila Badonnel were exposed to atmospheres containing 35% and 55% CO₂ for 30 generations to select strains resistant to high CO₂ content (HCC). Selection pressure was maintained at around 70% of mortality. At the 30th generation, comparison of sensitivities between the selected strains (HCC₁ and HCC₂) and the original susceptible strain (MA-S) revealed a resistance factor (RF) at the 50% mortality level (LT₅₀) of 4. 6-and 5. 3-folds, respectively. Throughout the selection process, log-time against probit-mortality lines remained roughly parallel and the slopes remained lower than that of MA-S strain until the last generation. It is inplied that the genetic potential of L. bostrychophila to develop resistance to CO₂ was not exhausted until the 30th generation. Removal of selection pressure for 5 generations from 2 sub-populations of two selected strains from the 25th generation caused significant reduction in resistance. In the absence of MAs exposure, the two selected strains (HCC₁ and HCC₂) possessed the fitness defect. HCC₁ and HCC₂ were calculated by R₀ to have a fitness value of 0. 52 and 0. 45 relative to MA-S.     

Discrimination between the regioisomeric 1,2- and 1,3-diacylglycerophosphocholines by phospholipases

The artificial 1,3-diacyl-glycero-2-phosphocholines (1,3-PCs), which form similar aggregate structures as the naturally occurring 1,2-diacyl-sn-glycero-3-phosphocholines (1,2-PCs), were tested as substrates for different classes of phospholipases such as phospholipase A₂ (PLA₂) from porcine pancreas, bee and snake venom, and Arabidopsis thaliana, phospholipase C (PLC) from Bacillus cereus, and phospholipase D (PLD) from cabbage and Streptomyces species. The regioisomers of the natural phospholipids were shown to bind to all investigated phospholipases with an affinity similar to the corresponding naturally occurring phospholipids, however their hydrolysis was reduced to different degrees (PLA₂s and PLC) or even abolished (PLDs belonging to the PLD superfamily). The results are in accordance with binding models obtained by docking the substrates to the crystal structures or homology models of the phospholipases.     

Screening and identification of epiphytic yeasts with potential for biological control of green mold of citrus fruits

Epiphytic yeasts isolated from the surface of citrus fruits, harvested in several orchards in the Souss-Massa-Draa Valley, Agadir, Morocco, were in vivo screened for antagonistic activity against Penicillium digitatum, the causal agent of green mold of citrus. From a total of 245 yeast strains assessed for their biocontrol activity against P. digitatum, fifteen reduced the incidence of disease to less than 50%. The effectiveness of the best selected yeast strains showed that Pichia anomala (YT73), Debaryomyces hansenii (YT22) and Hanseniaspora guilliermondii (YT13) were the most effective, with a reduction of green mold incidence from 65 to ~80%, compared to the control. The identification of the fifteen selected yeast strains was carried out through an integrated approach including phenotypic and genotypic (sequencing of D1/D2 domain of 26S rDNA encoding gene) methods. These 15 selected were identified as: H. guilliermondii, D. hansenii, H. uvarum and P. anomala. The study of the dynamics of two of the best strains, H. guilliermondii and D. hansenii, showed that these strains can grow rapidly, by approximately 2 log units, in citrus fruit wounds. Such rapid growth in wounds indicates that these antagonist yeasts are excellent colonizers of citrus wounds and can thrive on citrus fruits as a substrate.     

Further studies of the β-lactamases ofBacteroides species

The -lactamases of individual strains ofBacteroides fragilis, B. thetaiotaomicron, andB. melaninogenicus were examined to characterize their enzymatic activity and the relation between the periplasmic and cytoplasmic forms of the enzymes. K_m and V_max values indicate that all strains examined were very similar in terms of enzymatic activity with the antibiotics tested. Electrophoretic analysis and treatment with phospholipase D suggest the presence of a cytoplasmic form of the enzyme that is modified upon entry into the periplasmic space.     

海南地区与环渤海湾美人鱼发光杆菌美人鱼亚种水产动物分离株的表型与遗传特征分析

美人鱼发光杆菌美人鱼亚种(Photobacterium damselae subsp.damselae,PDD)是一种广泛分布于海洋环境内的重要病原菌,可导致多种海洋生物患病死亡,近年来在我国不同养殖区域内均有发现和报道.[目的]通过生理代谢表型、毒力基因分布和分子遗传分析,系统比较我国海南地区和环渤海湾不同株系PDD...     

Comparison of Some Photosynthetic Characters Between Two Hybrid Rice Combinations Differing in Yield Potential

Photosynthetic characteristics of two hybrid rice combinations, Peiai 64S/E32 and Shanyou 63, were compared at the panicle differentiation stage. As compared with Shanyou 63, the new combination Peiai 64S/E32 showed a significantly higher net photosynthetic rate (P _N), apparent quantum yield of carbon assimilation (_c), carboxylation efficiency (CE), and photorespiratory rate (R _P) as well as leaf chlorophyll content, but a significantly lower dark respiration rate (R _D) and compensation irradiance (I _c). It also showed a slightly higher photochemical efficiency (F_v/F_m and F/F_m) of photosystem 2, a lower non-photochemical quenching (q_N), and a similar CO₂ compensation concentration () as compared to Shanyou 63.     

The replicability of QTLs for murine alcohol preference drinking behavior across eight independent studies

On the basis of eight independent quantitative trait loci (QTL) studies of ethanol (alcohol) preference drinking in mice, a meta-analysis was carried out to examine the replicability of QTLs across studies and to enhance the power of QTL detection and parameter estimation. To avoid genetic heterogeneity, we analyzed only studies of mapping populations derived from the C57BL/6 (B6) and DBA/2 (D2) inbred progenitor strains. Because these studies were carried out in five different laboratories, there were substantial differences in testing procedure, data analysis, and especially in the choice of mapping population (BXD recombinant inbred strains, F₂, backcross, selected lines, or congenic strains). Despite this, we found several QTLs that were sufficiently robust as to appear consistently across studies given the strengths and weaknesses of the mapping populations employed. These were on Chromosomes (Chrs) 2 (proximal to mid), 3 (mid to distal), 4 (distal), and 9 (proximal to mid). The P value for each of these QTLs, combined across all applicable studies, ranged from 10⁻⁷ to 10⁻¹⁵, with the additive effect of each QTL accounting for 3–5% of the trait variance extrapolated to an F₂ population. Two other QTLs on Chrs 1 (distal) and 11 (mid) were less consistent, but still reached overall significance (P < .0001). Received: 18 April 2001 / Accepted: 25 July 2001     

Gastric body cholinergic contractile signal transduction in M2 and M3 receptor knockout mice

Abstract

Although most smooth muscles express a greater density of M₂ than M₃ muscarinic receptors, based on the potency of subtype selective muscarinic receptor antagonists, the M₃ subtype predominantly mediates contraction. The effect of inhibitors of putative contractile signal transduction pathway enzymes on carbachol-induced contractions was determined in wild-type (WT) mice and mice lacking either the M₂ (M₂KO) or the M₃ (M₃KO) receptor subtype. Contractile responses to KCl, then increasing carbachol concentrations in the presence and absence of enzyme inhibitors was determined. The KCl-induced contraction was not different between strains. The carbachol response was unaffected in the M₂KO strain but decreased 42% in M₃KO mice (p?<?0.01). Darifenacin potency was high in both WT and M₂KO strains, indicating M₃-mediated contractions, and low in the M₃KO strain, suggesting M₂-mediated contractions. The phosphatidyl inositol-specific phospholipase C (Pi-PLC) inhibitor ET-18-OCH₃ had no effect. Inhibition of phosphatidyl choline-specific phospholipase C (PC-PLC) and sphingomyelin synthase with D609 decreased maximal contraction in all strains. M₃-mediated contractions in the M₂KO strain were decreased 54% by the protein kinase C (PKC) inhibitor chelerythrine. M₂-mediated contractions in the M₃KO and WT strains were decreased by the Rho kinase (ROCK) inhibitor Y27632 as well as the ROCK, PKA and PKG inhibitor H89. The M₃ subtype activates PKC and either PC-PLC or sphingomyelin synthase, while the M₂ subtype activates ROCK and either PC-PLC or sphingomyelin synthase. These studies suggest that multiple parallel pathways mediate cholinergic contractions in stomach body smooth muscle.