Onset of penicillin-induced bacteriolysis in staphylococci is cell cycle dependent.

Synchronously growing staphylococci were treated with "lytic" concentrations of penicillin at different stages of their division cycle. Coulter Counter measurements and light microscopy were used to determine the onset of bacteriolysis. Independent of the stage of the division cycle at which penicillin was added, (i) the cells were always able to perform the next cell division; (ii) the following division, however, did not take place; and (iii) instead, at this time, when the onset of the subsequent cell separation was observed in control cultures, lysis of the penicillin-treated cells occurred. These results support a recent model (P. Giesbrecht, H. Labischinski, and J. Wecke, Arch. Microbiol. 141:315-324, 1985) explaining penicillin-induced bacteriolysis of staphylococci as the result of a special morphogenetic mistake during cross wall formation.     

Inactivation of recombinant plasmid DNA from a human erythropoietin-producing mouse cell line grown on a large scale

Summary Experiments were carried out to assess the survival of recombinant plasmid DNA during large-scale production of recombinant human erythropoietin (rhuEPO) in a fermentation pilot plant. The analyses revealed DNA-degrading activities in the fermentation broth and in the waste-water, leading to rapid destruction of plasmid DNA added to medium or waste-water. The capability of the plasmid-DNA-spiked samples to transform competent bacteria was drastically reduced. The DNA-degrading activity in the waste-waters could be blocked by addition of EDTA or by boiling, indicating the presence of DNA-degrading enzymes (DNases). No plasmid-specific DNA sequences were detected in waste-water samples by in-vitro amplification with Taqpolymerase. Genomic DNA preparations of cell debris collected from waste-water samples only contained degraded plasmid DNA. Furthermore, it was shown that intact plasmid DNA could be degraded to fragments of less than 1000 bp by incubation at 121°C for 20 min, leading to a decrease in the plasmid-specific transforming capacity by a factor of 10³ per minute. Thus, DNA from the rhuEPO production pilot plant was efficiently inactivated at three different levels: (i) in the fermentation medium (DNase), (ii) in the waste-water container (DNase), and (iii) by heat inactivation for 20 min at 120°C. These results indicate that the probability of delivery of recombinant DNA into the environment is extremely low in such biotechnological production processes. Offprint requests to: M. R. Fibi     

Chromosome abnormalities found among 34910 newborn children: results from a 13-year incidence study in Århus,Denmark

Summary As part of a larger prospective study of the influence of environmental factors on pregnancy, birth and the fetus, chromosome examinations have been made in 34910 newborn children in Århus over a 13-year period. Klinefelter's syndrome was found in 1 per 576 boys, XYY in 1 per 851 boys, triple-X in 1 per 947 girls and Turner's syndrome in 1 per 1893 girls. Other sex chromosome aberrations were found in 1 per 11637 children. The total incidence of sex chromosome abnormalities was 1 per 426 children or 2.34 per 1000. The most frequent autosomal abnormalities were that of Down's syndrome with 1 per 592 children, and reciprocal translocations with 1 per 712 children. The total incidence of autosomal abnormalities was 1 per 164 children. Chromosome abnormalities were found in 276 liveborn children and in 19 fetuses, who were aborted after prenatal chromosome examination. The combined incidence of sex chromosomal and autosomal abnormalities was 1 per 118 children or 8.45 per 1000 children.     

Glycine Antagonists Structurally Related to 4,5,6,7-Tetrahydroisoxazolo[5,4-c]pyridin-3-ol Inhibit Binding of [3H]Strychnine to Rat Brain Membranes

[3H]Strychnine binding to rat pons + medulla membranes was used as a measure of glycine receptors or glycine receptor-coupled chloride channels in vitro. A series of compounds structurally related to 4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol (THIP), which previously were shown to antagonize glycine responses in cat spinal cord, inhibited [3H]strychnine binding in micromolar concentrations. The most potent of these glycine antagonists, 5,6,7,8-tetrahydro-4H-isoxazolo[3,4-d]azepin-3-ol (iso-THAZ), was also the most potent inhibitor of [3H]strychnine binding, with a Ki of 1,400 nM. The Ki value for strychnine was 7.0 nM, whereas the Ki value for the mixed gamma-aminobutyric acid (GABA)/glycine antagonist 3 alpha-hydroxy-16-imino-5 beta-17-aza-androstan-11-one (RU 5135) was only 4.6 nM. Sodium chloride (1,000 mM) enhanced the affinity of strychnine, brucine, isostrychnine, and the nonselective GABA antagonist pitrazepin for [3H]strychnine binding sites, whereas the affinities of glycine, beta-alanine, and taurine were reduced. These sodium chloride shifts, however, were not predictive of antagonist or agonist properties, since the sodium chloride shift for the glycine antagonist iso-THAZ and of the other THIP-related antagonists were similar to those of the glycine-like agonists. The various sodium chloride shifts show that different groups of ligands bind to glycine receptor sites in different ways.     

Responses of aerial ventilation to hypoxia and hypercapnia inChanna argus,an air-breathing fish

Summary The snake-head fish (Channa argus) is an obligate air-breather inhabiting fresh waters in the temperate zone of East Asia.Ventilation of the air-breathing organ and aerial gas exchange were measured in 1 to 2 kg specimens at 15 and 25°C. Additionally, the ventilatory responses to hypoxia and hypercapnia were studied. Aerial ventilation increased from 1.1 to 2.9 mlbtps·kg^–1·min^–1 when temperature rose from 15 to 25°C. Concomitantly, O₂-uptake through airbreathing increased from 0.1 mlstpd·kg^–1·min^–1 (15°C) to 0.28 mlstpd·kg^–1·min^–1 (25°C), whereas aerial gas exchange was less important for CO₂-climination as evident from low gas exchange ratios (0.16 at 15°C, 0.29 at 25°C).Ventilation increases only slightly in response to inspiration of hypercapnic gas mixtures or to hypoxic conditions in water. By contrast, inspiration of hypoxic gas mixtures caused marked increases of ventilation in particular at the higher temperature.Aerial ventilation inChanna is low compared to values for ectothermic pulmonary breathers. However, its ventilatory responses to hypoxia strikingly resemble those of reptiles: The most marked ventilatory response to hypoxia occurs at the higher temperature where the demands for O₂ are greatest.     

The quantification of the haemagglutinin content of influenza whole virus and Tween-ether split vaccines

Monovalent whole virus and Tween-ether split vaccines prepared from influenza A/Bangkok, A/Brazil and B/Singapore were assayed for haemagglutinin content using single radial immunodiffusion (SRID), quantitative sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunization of guinea pigs. When SRID was performed with split vaccines, haemagglutinin values were consistently recorded which were in the range of 50 to 25% of the values obtained before disruption of virions. When, however, disruption was conducted in the presence of excess detergent, thus preventing aggregate formation of solubilized haemagglutinin, test values comparable with those of whole virus vaccines were obtained. In agreement with these results, immunization experiments revealed that whole virus and corresponding split vaccines exhibited comparable immunogenicity in guinea pigs. Additionally it could be calculated from SDS-PAGE and densitometer tracings, obtained by scanning the gels after staining with either Coomassie blue or FITC-Con A, that 90 to 95% of whole virus HA2 was recovered in Tween-ether split vaccines. On the basis of these findings we conclude that precise quantification of Tween-ether split vaccines is not possible by the SRID test alone. As aggregate formation of solubilized haemagglutinin occurs, we suggest that either a physico-chemical method including a disaggregation procedure, such as SDS treatment, or immunological evaluation of the original whole virus preparation before disruption of virions should be applied as an additional criterion for quantification of influenza Tween-ether split vaccines.     

Rates of Production of Individual Volatile Fatty Acids in the Rumen of Lactating Cows

The rumen fermentation rates in individual lactating cows were measured in four different experiments. The results disclosed that the amounts and proportions of volatile acids formed could vary widely. In one case, a marked difference in the proportions of the acids produced arose within the experiment and correlated with a difference in the proportion of methane formed.

The average rate of production per day was 10.5 moles butyric acid, 12.8 moles propionic acid, and 40 moles acetic acid. Manometric estimations of rate gave lower results than those obtained by the zero-time method, due to delay after sampling and to failure of the acids to liberate stoichiometric quantities of carbon dioxide.

For those experiments in which zero-time rates were estimated, the average specific absorption rates, i.e., the amount absorbed per hour per micromole of acid in the rumen, were 0.37 for butyric acid, 0.38 for propionic acid, and 0.26 for acetic acid.

The carbon dioxide, acids, and microbial cells produced in the rumen fermentation are estimated to account for about 90% of the carbon found in the milk and respiratory CO₂ of the cows. The carbon dioxide from the fermentation was about 27% of the carbon dioxide exhaled.

     

Predation and potential transfer of pollen in a population of Saxifraga hirculus

Body surface pollen load and gut pollen (pollen diet load) of flower visitors in a Danish population of Saxifraga hirculus were analyzed. The four most frequent visitors, Eurimyia lineata (Syrphidae), Neoascia tenur (Syrphidae), Asindulum nigrum (Mycetophilidae), and Zygaena trifolii (Zygaenidae) had total surface loads (and S. hirculus loads) of 1031 (771), 137 (106), 108 (78), and 318 (27) pollen grains, respectively. Other syrphid visitors had on average a load of 407 (192) pollen grains. Males of E. lineata, N. tenur , and A. nigrum carried more surface pollen than the females; females of E. lineata had a lot of pollen from Ranunculus acris and S. hirculus and males had a lot from S. hirculus . Females of N. tenur carried pollen mostly from S. hirculus, Lotus uliginosus and Galium uliginosum ; males carried pollen mostly from S. hirculus . The surface pollen load of A. nigrum was dominated by S. hirculus pollen, whereas that of Z. trifolii was dominated by Cirsium palustre and Lychnis flos-cuculi pollen. The amount and kind of pollen varied enormously both intraspecifically and interspecifically. Eurimyia lineata, N. tenur and other syrphids had total pollen diet loads (and S. hirculus pollen diet loads) of approximately 14900 (657), 3739 (2327) and 17784 (13040). The average syrphid contained pollen from five plant species, only one to two, however, were quantitatively important. The average diet load in females was nine times that in males in both E. lineata and N. tenur. Eurimyia lineata ate a lot of R. acris; N. tenur ate a lot of S. hirculus and G. uliginosum . Generally, no correlation was observed between size or season and the composition of surface load and diet load. Eurimyia lineata made most of the pollen visits. Thus, the seed set from this Danish population of S. hirculus was dependent on a single syrphid species.     

Evidence for the Colonization of Lactic Acid Bacteria in the Gastrointestinal Tract of Suckling Mink

Lactic acid bacteria are considered indigenous members of the gastrointestinal microflora in a number of animal species (Savage 1977a). Some intestinal strains of lactobacilli and streptococci are aWe to adhere to stratified squamous epithelium of some animals (Tannock et al. 1987), in the non-secreting part of the stomach of piglets (Barrow et al. 1980, Fuller et al. 1978) and rodents (Tannock et al. 1982), and in the crop of poultry (Fuller 1978). The presence of lactic acid bacteria in the digestive tract is believed to be of beneficial value to the host animal (Fuller 1989). The production of organic acids in the stomach or the crop helps maintaining a low pH which may be important for inhibiting the colonization of potentially pathogenic bacteria, particularly in the newborn animal (Barrow et al 1980, Fuller 1977, Fuller 1978). The adhesion of lactobacilli to squamous epithelium is host specific: strains capable of adhering to the epithelium of piglets are usually not able to adhere in rodents or poultry and vice versa (Fuller 1978, Lin & Savage 1984, Tannock et al 1982). Adhesion of lactic acid bacterial strains to other epithelia than stratified squamous epithelium has been reported. Thus, the attachment of lactobacilli to cells from the secreting epithelium of the murine stomach (Kotarski & Savage 1979), to intestinal cells of humans (Goldin & Gorbach 1987), and to columnar epithelial cells of piglets and calves (Mäyrä-Mäkinen et al 1983) has been demonstrated using in vitro methods. In another study the in vivo attachment of Enterococcus faecium to duodenal epithelium of gnotobi-otic chickens was demonstrated (Fuller et al 1981). Recent research indicated that in adult mink lactic acid bacteria are not indigenous members of the intestinal flora, and they do not attach to epithelium in any part of the gastrointestinal tract (Federsen & Jørgensen 1992). The present paper presents evidence that Gram positive cocci may colonize the gut of suckling mink kits and attach to the gut mucosa.     

Technetium and rhenium complexes of mercapto-containing peptides 1. Tc(V) and Re(V) complexes with mercaptoacetyl diglycine (MAG2) and X-ray structure of AsPh4[TcO(MAG2)]·C2H5OH

The reaction of mercaptoacetyl diglycine (MAG₂) with technetium(V) gluconate in aqueous solution produced [TcO(MAG₂)]⁻. A single X-ray structure determination was carried out for the tetraphenylarsonium salt. The dark brown crystals are monoclinic, space group P2(1)/n, with a=12.478(5), b=14.922(5), c=17.183(9) Å and Z=4. The [TcO(MAG₂)]⁻ ion has a square pyramidal geometry with the technetium atom displaced by 0.756 Å towards the oxo ligand from the plane formed by the equatorial S,N,N,O atoms. The rhenium complex AsPh₄[ReO(MAG₂)] was prepared analogously starting from Re(V) gluconate and characterized.