Stability and conjugal transfer kinetics of a TOL plasmid in Pseudomonas aeruginosa PAO 1162

Abstract Batch mating experiments were employed to study the kinetics of the conjugal transfer of a TOL plasmid, using the transconjugant strain Pseudomonas aeruginosa PAO 1162 (TOL) as the plasmid donor and Pseudomonas putida PB 2442 and Pseudomonas aeruginosa PAO 1162N as the plasmid recipients. Transfer rates from PAO 1162 (TOL) to PAO 1162N and PB 2442 measured for exponentially grown PAO 1162 (TOL) were 1.81 × 10⁻¹⁴ (standard error (S.E.) 1.25 × 10⁻¹⁵) ml·cell⁻¹min⁻¹ and 3.32 × 10⁻¹³ (S.E. 4.42 × 10⁻¹⁴) ml·cell⁻¹min⁻¹, respectively. The instability of the TOL plasmid in PAO 1162 (TOL) was evaluated under conditions that were non-selective for maintenance of the TOL catabolic functions. The measured rates of instability were 6.7 10⁻⁶ to 8.3 10⁻⁶ min⁻¹, and the loss of the catabolic functions was mainly caused by structural instability of the plasmid.     

Cell-surface properties of enterotoxigenic and cytotoxic Salmonella enteritidis and Salmonella typhimurium: studies on hemagglutination, cell-surface hydrophobicity, attachment to human intestinal cells and fibronectin-binding

Thirteen Salmonella enteritidis and S. typhimurium strains with smooth or rough colony morphology were investigated for their surface properties based on hemagglutination (HA), hydrophobicity, and fibronectin-binding profiles. The strains showed 5 different patterns of HA which was mannose-sensitive. The rough strains possessed comparatively greater number of fimbriae than the corresponding smooth strains and also attached to human intestinal cells in greater numbers. The Salmonella strains used in this study interacted with fibronectin and its 29-kDa N-terminal fragment to varied extents. These properties may be helpful in broadening the prospective interaction capabilities of Salmonella organisms with the host surfaces.     

A light intensity threshold for schooling in the Atlantic mackerel, Scomber scombrus

The schooling behaviour of Atlantic mackerel was studied in a large tank at different light intensities in the range 12.6–1.8 × 10⁻¹⁰μEs⁻¹ m⁻². Variable light intensity was produced by accurately controlling the current to a green light-emitting diode (LED) 3 m above the experimental tank. Under high light levels (1.8 × 10⁻⁶μEs⁻¹ m⁻²) mackerel always formed a single school, whereas at lower levels (1.8 × 10⁻⁸μEs⁻¹ m⁻²) they swam as individuals. At light levels down to 1.0 × 10⁻⁶μEs⁻¹ m⁻² the mean nearest neighbour distance in a school remained relatively constant (0.3–0.9 body lengths), and individual mackerel swam along a path which deviated from the position of their nearest neighbours by less than 14°. As light dropped below 1.8 × 10⁻⁷μEs⁻¹ m⁻², both nearest neighbour distance and heading angle between nearest neighbours increased, with mean values of 1–1.8 body lengths and 23–92°, respectively, at 1.8 × 10⁻⁹μEs⁻¹ m⁻². The results are discussed in terms of ambient light conditions in the sea.     

Quinuclidinyl Benzilate Binding in House Fly Heads and Rat Brain

Abstract: House fly heads contain a binding site for 3-quinuclidinyl benzilate (QNB) that is quite similar in pharmacology to the muscarinic acetylcholine receptor of vertebrate tissues. The house fly site binds [³H]QNB reversibly with a K _d of 260 PM and B_max of 1 pmol/g of heads from direct binding measurements. The K_d calculated from the ratio of the dissociation rate constant (2 × 10⁻⁴ sec⁻¹) to the association rate constant (2.5 × 10⁶ M⁻¹ Sec⁻¹) was 80 pM. The house fly site binds (-)quinuclidinyl benzilate preferentially, as do classic muscarinic receptors. The binding is also sensitive to other muscarinic antagonists and agonists. Nicotinic and other drugs are no more effective on the house fly site than they are on the rat brain muscarinic receptor itself. These binding studies suggest that the house fly QNB binding site is a muscarinic receptor.     

The variant rpoS allele of S. enteritidis strain 27655R does not affect virulence in a chick model nor constitutive curliation but does generate a cold-sensitive phenotype

Adherence of pathogenic Escherichia coli and Salmonella spp. to host cells is in part mediated by curli fimbriae which, along with other virulence determinants, are positively regulated by RpoS. Interested in the role and regulation of curli (SEF17) fimbriae of Salmonella enteritidis in poultry infection, we tested the virulence of naturally occurring S. enteritidis PT4 strains 27655R and 27655S which displayed constitutive and null expression of curli (SEF17) fimbriae, respectively, in a chick invasion assay and analysed their rpoS alleles. Both strains were shown to be equally invasive and as invasive as a wild-type phage type 4 strain and an isogenic derivative defective for the elaboration of curli. We showed that the rpoS allele of 27655S was intact even though this strain was non-curliated and we confirmed that a S. enteritidis rpoS::str^r null mutant was unable to express curli, as anticipated. Strain 27655R, constitutively curliated, possessed a frameshift mutation at position 697 of the rpoS coding sequence which resulted in a truncated product and remained curliated even when transduced to rpoS::str^r. Additionally, rpoS mutants are known to be cold-sensitive, a phenotype confirmed for strain 27655R. Collectively, these data indicated that curliation was not a significant factor for pathogenesis of S. enteritidis in this model and that curliation of strains 27655R and 27655S was independent of RpoS. Significantly, strain 27655R possessed a defective rpoS allele and remained virulent. Here was evidence that supported the concept that different naturally occurring rpoS alleles may generate varying virulence phenotypic traits.     

Conjugative transfer of the transposon Tn919 to lactic acid bacteria

Abstract The streptococcal transposon Tn 919 was transferred from Streptococcus faecalis GF590 to selected Group N Streptococcus strains and to one strain each of Lactobacillus plantarum and Leuconostoc cremoris , using the filter mating method. An S. lactis MG1363 Rif^r Tc^r transconjugant also acted as a donor, but was less efficient than GF590. Frequencies of transfer varied between 4.0 × 10⁻⁸ and 5.29 × 10⁻⁵ per recipient. Further analysis of S. lactis MG1363 Sm^r Tc^r transconjugants showed that insertion of Tn 919 into the chromosome was site-specific.     

Field estimates of oxygen consumption for the brine shrimp Parartemia zietziana Sayce (Crustacea: Anostraca) in two salt lakes in Victoria, Australia

SUMMARY. Oxygen consumption of P. zietziana was measured monthly in two saline (>60‰ salinity) lakes from November 1973 to November 1975 with short (<2 h) in situ incubations in BOD bottles. Tests in which oxygen decline was monitored continuously showed that there was no handling effect and respiratory rate was constant down to 1.8–1.9 mg O₂ 1⁻¹, about 40% of the usual initial concentration. Incubations over 24 h demonstrated no diurnal fluctuations in oxygen consumption. Multiple regression analysis indicated that 90% of the variance in respiratory rate ( R in mg O₂x10⁻⁴h⁻¹ individual⁻¹) was accounted for by changes in salinity (3%; S in ‰), temperature (7%; T in °C) and dry weight (8%; W in mg × 10⁻³): log R =−1.123+0.0025+0.021 T+ 0.756 log W. From this equation and data on population density, population respiration was calculated: 91864.5 mg O₂ m⁻² year⁻¹ in Pink Lake and 12367.5 mg O₂ m⁻²year⁻¹ in Lake Cundare.     

Induction of a wheat Em promoter by ABA and optically pure ABA analogs in white spruce (Picea glauca) protoplasts

In white spruce ( Picea glauca ) protoplasts, abscisic acid (ABA) and optically pure ABA analogs induced expression of a reporter gene under regulation of a wheat ABA-responsive promoter. A fusion of a 650 bp promoter fragment from the wheat Em gene promoter and the Escherichia coli uidA sequence encoding β -glucuronidase (GUS) was linked in the plasmid pBM 113Kp. Expression of the Em-uidA fusion varied among 6 white spruce genotypes. Protoplasts from 4-day-old embryogenic suspension cultures gave the highest GUS activity relative 10 other stages in the 7-day growth cycle of suspension cultures. Racemic ABA [R.S-(±)-ABA] induced a significant increase of protoplast GUS activity over background at a concentration of 1 × 10⁻⁵ M , but maximum GUS activity was found at 1 × 10⁻³ M , ABA stereochemistry had a significant effect on gene expression. The natural isomer of ABA [S-(+)-ABA] was an effective inducer at a concentration as low as 1 × 10⁻⁷ M , but a concentration of greater than 1 × 10⁻⁴ M was required for induction by [R-(—)-ABA]. Moreover, analogs with the same configuration at C-l¹ as that of natural ABA were more effective for induction of expression from the Em-uidA . insert at 1 × 10⁻⁴ M than were their enamiomers. Plasnud pBI511. carrying the chloramphenicol acety] transferase (CAT) gene driven by the constitutively expressed, tandemly duplicated cauliflower mosaic virus 35S promoter, was co-electroporated with pBM113Kp for monitoring Ihe influence of addition of exogenous ABA or ABA analogs on heterologous gene expression in protoplasts. CAT activity was not significantly affected by the presence or absence of ABA or the analogs used.     

Helicobacter pylori interacts with heparin and heparin-dependent growth factors

Abstract The pathogenic bacterium Helicobacter pylori , which causes active, chronic type B gastritis and peptic ulcer disease, and increases the risk for development of gastric cancer, could tentatively interfere with growth factors and growth factor receptors of importance for the gastroduodenal mucosa, e.g. heparin-binding FGFs (fibroblast growth factors). H. pylori binds FGF with an extremely strong affinity (3.8 × 10⁻¹² M), and also heparan sulfate and heparin with higher affinity ( K _d 9 × 10⁻⁹ M) than FGFs bind to heparin (10⁻⁸–10⁻⁹ M). FGF receptors are also dependent on heparin for their activation. Heparan sulfate binding proteins (HSBP) are exposed on and shed from the surface of H. pylori , which often are localised close to the epithelial stem cells in the gastroduodenal glands. H. pylori could thus efficiently interfere with growth factors and growth factor receptors, tentatively resulting in disturbance of the delicate balance that control the renewal, maintenance and repair of the gastroduodenal mucosa. This mode of action has previously not been considered, but may constitute part of its pathogenic mechanism. Such a dynamic mode of action of H. pylori may explain the reason for that infected victims may either suffer from gastrointestinal symptoms or lack clinical evidence of disease or discomfort.     

Detection of the response regulator AgrA in the cytosolic fraction of Staphylococcus aureus by monoclonal antibodies

Abstract The interaction of salivary lysozyme with the surface protein antigen (PAc) of Streptococcus mutans and the interaction of lysozyme with the pathogen were examined by ELISA using S. mutans MT8148 (PAc⁺) and the PAc-defective mutant EM-2 (PAc⁻). The lysozyme clearly bound to the S. mutans wild type but not to the S. mutans mutant. Furthermore, lysozyme bound directly in the fluid phase to the rPAc, of which the binding kinetics were determined ( K _on= 3.63 ± 0.04 × 10³M⁻¹ s ⁻¹, K _off= 1.72 ± 0.04 × 10⁻⁵s⁻¹ and K_on / K_off= 2.11 × 10⁸M⁻¹) using surface plasmon resonance. The kinetics of both association and dissociation were relatively slow. In addition, anti-lysozyme antibody significantly inhibited the binding of salivary components to the rPAc. The present findings indicate that lysozyme is one of the major salivary components interacting with S. mutans PAc.     

Uptake and metabolism of taurine in the green alga Chlorella fusca

Taurine entered the alga Chlorella fusca Shihira et Krauss strain 21l-8b via a pH and energy-dependent system ("permease"). Transport followed triphasic kinetics from 10⁻⁶ to 10⁻² M with K_m values for taurine of 5.4 × 10⁻⁵, 4.1 × l0⁻⁴ and l.5 × 10⁻³ M. This uptake system was specific for sulfonic acids and showed no affinity for α- and β -amino acids or Na⁺; thus the permease of C. fusca is different from all known taurine transport systems with respect to structural specificity and lack of Na⁺ -dependence. Uptake was not observed in sulfate-grown algae but developed as a response to sulfate limitation within 2 h. Sulfate addition caused a rapid decline in taurine transport capacity. Labeled taurine was rapidly metabolized in C. fusca to sulfate and ethanolamine, suggesting oxidative hydrolysis as the mechanism of C-S bond cleavage. Further incorporation of these catabolic products in C - and S -metabolism was demonstrated. Taurine catabolism was also detected in other green algae and some cyanobacteria.     

Introduction of the mercury transposon Tn501 into Rhizobium japonicum strains 31 and 110

Abstract Transposon Tn 501 , which encodes resistance to mercuric ions, was introduced into Rhizobium japonicum 110 and 31 by conjugal transfer. The transposon donor plasmid (pMD100) was able to mobilize into R. japonicum , but could not be maintained. Hg²⁺-resistant colonies were recovered at a frequency of 1.9 × 10⁻⁸/recipient for strain 110, and 1.7 × 10⁻⁷/recipient for strain 31. Presence of Tn 501 in Hg-resistant isolates was verified by Southern analysis and demonstrating transposition of Hg resistance. Transposon mutagenesis has been used to generate auxotrophic mutations at low frequency.     

Enzymes involved in ammonia assimilation in the fungus Acremonium persicinum

Abstract Acremonium persicinum grown in batch culture with ammonium tartrate as the nitrogen source possessed an NADP⁺-dependent glutamate dehydrogenase and a glutamine synthetase. Glutamate synthase was not detected under the culture conditions used. Kinetic studies of the NADP⁺-dependent glutamate dehydrogenase at 25°C and pH 7.6 revealed an apparent K _m of 3.2 × 10⁻⁴ M for 2-oxoglutarate and an apparent K _m of 1.0 × 10⁻⁵ M for ammonium ions, with corresponding apparent V _max values of 0.089 and 0.13 μmol substrate converted/min/mg of protein, respectively. Glutamine synthetase was measured by the γ-glutamyl transferase reaction at 30°C and pH 7.55. This transferase reaction of glutamine synthetase had a higher rate at 30°C than at 25°C or 37°C.     

FIRST REPORT OF THE CYANOTOXINS CYLINDROSPERMOPSIN AND DEOXYCYLINDROSPERMOPSIN FROM RAPHIDIOPSIS CURVATA (CYANOBACTERIA)

A strain of Raphidiopsis (Cyanobacteria) isolated from a fish pond in Wuhan, P. R. China was examined for its taxonomy and production of the alkaloidal hepatotoxins cylindrospermopsin (CYN) and deoxy-cylindrospermopsin (deoxy-CYN). Strain HB1 was identified as R. curvata Fritsch et Rich based on morphological examination of the laboratory culture. HB1 produced mainly deoxy-CYN at a concentration of 1.3 mg·g ^{− 1} (dry wt cells) by HPLC and HPLC-MS/MS. CYN was also detected in trace amounts (0.56 μg·g ^{− 1}). A mouse bioassay did not show lethal toxicity when tested at doses up to 1500 mg dry weight cells·kg ^{− 1} body weight within 96 h, demonstrating that production of primarily deoxy-CYN does not lead to significant mouse toxicity by strain HB1. The presence of deoxy-CYN and CYN in R. curvata suggests that Raphidiopsis belongs to the Nostocaceae, but this requires confirmation by molecular systematic studies. Production of these cyanotoxins by Raphidiopsis adds another genus, in addition to Cylindrospermopsis , Aphanizomenon , and Umezakia , now known to produce this group of hepatotoxic cyanotoxins. This is also the first report from China of a CYN and deoxy-CYN producing cyanobacterium.     

Photosensitivity of Rumex obtusifolius seeds for stimulation of germination: Influence of light and temperature

A population of Rumex obtusifolius L. seeds imbibed for 24 h at 25°C exhibits a sigmoid logarithmic fluence-response relationship for stimulation of germination by red light (R), 11.0 μmol m⁻² being necessary for 50% of the response. After 24 h imbibition at 35°C the fluence-response relationship for stimulation of germination by R is biphasic. For 50% response the very sensitive phase (very low fluence-response) requires 4.7 − 10⁻²μmol m⁻² whereas the less sensitive phase (low fluence-response) requires 4.0 μmol m². A few seconds of far-red light (FR) satisfies the germination requirement of the sensitive seeds after 24 h at 35°C. However, a longer period of FR (2 h) results in low germination. The fluence-response relationship for induction of these seeds by R is sigmoid, 4.8 μmol m⁻² being necessary for 50% response, demonstrating that 2 h FR desensitizes the sensitive proportion of the seed population induced by 24 h at 35°C. A proportion of the seed population can be further sensitized by 60 min at 35°C following this desensitization.     

Identification of Strychnine Binding Sites in the Rat Retina

Abstract: [³H]Strychnine specifically binds to membrane fractions isolated from rat retinae. The binding is saturable, with an apparent dissociation constant, K _D, of 14.3 × 10⁻⁹ M and 205 fmol bound/mg protein. Specific binding is time-dependent and proportional to protein concentration. Glycine and taurine are equally potent inhibitors of [³H]strychnine binding ( K _i= 4 × 10⁻⁵ M); no other amino acids endogenously present in the retina inhibited [³H]strychnine binding.     

Permeability of the Chara cell membrane for ethylene glycol, glycerol, meso-erythritol, xylitol and mannitol

The permeability of internodal cells of Chara australis R. Brown for polyol molecules was determined by using a turgor balance to measure the increase in the osmotic pressure of an internodal cell incubated in artificial pond water containing one of the polyol compounds tested. The permeabilities for ethylene glycol, glycerol, meso -erythritol, xylitol and mannitol were (4. 39 ± 0. 52) × 10⁻⁹, (1. 49 ± 0. 40) × 10¹⁰, (4. 92 ± 0. 27) × 10⁻¹⁰ (9. 9 ± 3. 4) × 10¹¹ and (7. 6 ± 4. 8) × 10⁻¹² m s⁻³, respectively. The permeability for glycerol was slightly smaller than that for meso -erythritol, whose molecular weight is larger than that of glycerol in this homologous series: but the reason for this is not clear.     

Demethylation and cleavage of dimethylsulfoniopropionate in marine intertidal sediments

Abstract Demethylation and cleavage of dimethylsulfoniopropionate (DMSP) was measured in three different types of intertidal marine sediments: a cyanobacterial mat, a diatom-covered tidal flat and a carbonate sediment. Consumption rates of added DMSP were highest in cyanobacterial mat slurries (59 μmol DMSP 1⁻¹) and lower in slurries from a diatom mat and a carbonate tidal sediment (24 and 9 μmol DMSP 1⁻¹ h⁻¹, respectively). Dimethyl sulfide (DMS) and 3-mercaptopropionate (MPA) were produced simultaneously during DMSP consumption, indicating that cleavage and demethylation occurred at the same time. Viable counts of DMSP-utilizing bacteria revealed a population of 2 × 10⁷ cells cm⁻³ sediment (90% of these cleaved DMSP to DMS, 10% demethylated DMSP to MPA) in the cyanobacterial mat, 7 × 10⁵ cells cm⁻³ in the diatom mat (23% cleavers, 77% demethylators), and 9 × 10⁴ cells cm⁻³ (20% cleavers and 80% demethylators) in the carbonate sediment. In slurries of the diatom mat, the rate of MPA production from added 3-methiolpropionate (MMPA) was 50% of the rate of MPA formation from DMSP. The presence of a large population of demethylating bacteria and the production of MPA from DMSP suggest that the demethylation pathway, in addition to cleavage, contributes significantly to DMSP consumption in coastal sediments.     

Sublethal effects of imidacloprid and pymetrozine on population growth parameters of cabbage aphid, Brevicoryne brassicae on rapeseed, Brassica napus L.

Efficiency of imidacloprid and pymetrozine on population growth parameters of cabbage aphid, Brevicoryne brassicae L. （Homoptera： Aphididae） was determined using demographic toxicology by leaf dip method. At first, bioassay tests were performed. The LC50 value and confidence limit for imidacloprid and pymetrozine were 1.61 × 10^-5 mol/L （0.74 × 10^-5-2.66 × 10^-5） and 2.14 × 10^-4 mol/L （1.24 × 10^-4-3.40 × 10^-4）, respectively. To evaluate the sublethal effect of two insecticides on population growth parameters of cabbage aphid, LC30 concentrations of imidacloprid and pymetrozine were used at 5 mol/L and 30 mol/L. The experiments were carried out in a incubator at 20±1℃, 60% ± 5% RH and 16：8 （L：D） photoperiod on canola seedlings, Brassica napus L. var. ＇PF＇. Net fecundity rate decreased in both insecticide-treated populations. Intrinsic rates of increase （rm） were lower in imidacloprid and pymetrozine treatments than in controls. Intrinsic birth rates also decreased in treated populations. There was a relative increase in intrinsic death rates of treated populations. The mean generation times and doubling time were also lower in populations treated with insecticides than in controls. There was a considerable reduction in the average numbers of nymphs reproduced per female as compared with the control. The average longevity of female adults in the control was significantly different from those treated with imidacloprid and pymetrozine. However, there was no significant differences in aphid life-table parameters between the two insecticide-treated populations （P 〉 0.01）.     

Apoplastic mobility of sucrose in storage parenchyma of sugar beet

The apoplastic movement of sucrose through storage parenchyma discs (2.4 mm thick) from roots of sugar beet ( Beta vulgaris var. altissima ) was investigated in order to evaluate the suitability of the apoplast for transcellular sugar transport. The sucrose permeability of the discs (P = 5.7 × 10⁻⁸ cm s⁻¹ at 25°C) was more than two orders of magnitude lower than that of an equally thick layer of unstirred water. This is due to the small volume fraction of free space (3.1%) and the decreased diffusion coefficient D of sucrose in the cell walls. The effective diffusion coefficient of the apoplast (6 to 9 × 10⁻⁷ cm² s⁻¹ at 25°C) was determined independently of the cross sectional area of free space by treating the time course of fluxes according to Fick's second law. The high diffusion resistance of the apoplast has to be considered in models of native parenchyma transport.