Presence and Analysis of Plasmids in Human and Animal Associated Arcobacter Species

In this study, we report the screening of four Arcobacter species for the presence of small and large plasmids. Plasmids were present in 9.9% of the 273 examined strains. One Arcobacter cryaerophilus and four Arcobacter butzleri plasmids were selected for further sequencing. The size of three small plasmids isolated from A. butzleri and the one from A. cryaerophilus strains ranged between 4.8 and 5.1 kb, and the size of the large plasmid, isolated from A. butzleri, was 27.4 kbp. The G+C content of all plasmids ranged between 25.4% and 26.2%. A total of 95% of the large plasmid sequence represents coding information, which contrasts to the 20 to 30% for the small plasmids. Some of the open reading frames showed a high homology to putative conserved domains found in other related organisms, such as replication, mobilization and genes involved in type IV secretion system. The large plasmid carried 35 coding sequences, including seven genes in a contiguous region of 11.6 kbp that encodes an orthologous type IV secretion system found in the Wolinella succinogenes genome, Helicobacter pylori and Campylobacter jejuni plasmids, which makes this plasmid interesting for further exploration.     

Multiple Bacteriocin Production by Leuconostoc mesenteroidesTA33a and Other Leuconostoc/Weissella Strains

Leuconostoc (Lc.) mesenteroides TA33a produced three bacteriocins with different inhibitory activity spectra. Bacteriocins were purified by adsorption/desorption from producer cells and reverse phase high-performance liquid chromatography. Leucocin C-TA33a, a novel bacteriocin with a predicted molecular mass of 4598 Da, inhibited Listeria and other lactic acid bacteria (LAB). Leucocin B-TA33a has a predicted molecular mass of 3466 Da, with activity against Leuconostoc/Weissella (W.) strains, and appears similar to mesenterocin 52B and dextranicin 24, while leucocin A-TA33a, which also inhibited Listeria and other LAB strains, is identical to leucocin A-UAL 187. A survey of other known bacteriocin-producing Leuconostoc/Weissella strains for the presence of the three different bacteriocins revealed that production of leucocin A-, B- and C-type bacteriocins was widespread. Lc. carnosum LA54a, W. paramesenteroides LA7a, and Lc. gelidum UAL 187-22 produced all three bacteriocins, whereas W. paramesenteroides OX and Lc. carnosum TA11a produced only leucocin A- and B-type bacteriocins. Received: 11 April 1997 / Accepted: 10 June 1997     

Pollen–plant–climate relationships in sub-Saharan Africa

Aim  To demonstrate that incorporating the bioclimatic range of possible contributor plants leads to improved accuracy in interpreting the palaeoclimatic record of taxonomically complex pollen types.
Location  North Tropical Africa.
Methods  The geographical ranges of selected African plants were extracted from the literature and geo-referenced. These plant ranges were compared with the pollen percentages obtained from a network of surface sediments. Climate-response surfaces were graphed for each pollen taxon and each corresponding plant species.
Results  Several patterns can be identified, including taxa for which the pollen and plant distributions coincide, and others where the range limits diverge. Some pollen types display a reduced climate range compared with that of the corresponding plant species, due to low pollen production and/or dispersal. For other taxa, corresponding to high pollen producers such as pioneer taxa, pollen types display a larger climatic envelope than that of the corresponding plants. The number of species contained in a pollen taxon is an important factor, as the botanical species included in a taxon may have different geographical and climate distributions.
Main conclusions  The comparison between pollen and plant distributions is an essential step towards more precise vegetation and climate reconstructions in Africa, as it identifies taxa that have a high correspondence between pollen and plant distribution patterns. Our method is a useful tool to reassess biome reconstructions in Africa and to characterize accurately the vegetation and climate conditions at a regional scale, from pollen data.     

Retinoic acid synthesis by cytosol from the alcohol dehydrogenase negative deermouse

Cytosolic alcohol dehydrogenase in the deermouse is coded by a single genetic locus and a strain of the deermouse which is alcohol dehydrogenase negative exists. These two strains of the deermouse were used to extend insight into the role of cytosolic alcohol dehydrogenases in the conversion of retinol into retinoic acid. Retinoic acid synthesis from physiological concentrations of retinol (7.5 microM) with cytosol from the alcohol dehydrogenase negative deermouse was 13% (liver), 14% (kidney), 60% (testes), 78% (lung), and 100% (small intestinal mucosa) of that observed with cytosol from the positive deermouse. The rates in the negative strain ranged from 0.3 to 0.7 nmol/h/mg protein: sufficient to fulfill cellular needs for retinoic acid. Ten millimolar 4-methylpyrazole inhibited retinoic acid synthesis 92, 94, 26, and 30% in kidney, liver, lung, and testes of the positive deermouse, respectively, but only 50, 30, 0, and 0% in the same tissues from the negative deermouse. Ethanol (300 mM) did not inhibit retinoic acid synthesis in kidney cytosol from the negative strain. Therefore multiple cytosolic dehydrogenases, including alcohol dehydrogenases, contribute to retinol metabolism in vitro. The only enzyme(s) likely to be physiologically significant to retinoic acid synthesis in vivo, however, is the class of dehydrogenase, distinct from ethanol dehydrogenase, that is common to both the positive and the negative deermouse. This conclusion is supported by the data described above, the kinetics of retinoic acid synthesis and retinal reduction in kidney cytosol from the negative deermouse, and the very existence of the alcohol dehydrogenase negative deermouse. This work also shows that microsomes inhibit the cytosolic conversion of retinol into retinoic acid and that the synthesis of retinal, a retinoid that has no known function outside of the eye, does not reflect the ability or capacity of a sample to synthesize retinoic acid.     

Attempt to detect recombination between B-F and B-L genes within the chicken B complex by serological typing,in vitro MLR,and RFLP analyses

In search for recombinants within the chicken major histocompatibility B complex, 1155 animals from crosses between the congenic lines CB (B¹²) and CC (B⁴) were tested with alloantibodies and monoclonal antibodies for the B-F (class I), B-L (class II), and B-G (class IV) antigens and by mixed lymphocyte reaction. The absence of detectable recombination was confirmed by restriction fragment length polymorphism analysis with B-L and B-F probes. Together with previous reports, this indicates that the distance between the B-F and B-L loci is below 0.01 centimorgan.     

Sib pair linkage analysis of renin gene haplotypes in human essential hypertension

Summary Although essential arterial hypertension is believed to have a strong genetic predisposition, the gene(s) responsible are unknown. The mechanisms underlying the regulation of blood pressure and experimental studies place the renin gene among the main candidate genes that need to be tested in humans. We tested the hypothesis of a linkage between the renin gene and essential hypertension using the affected sib pair method. Siblings (133 subjects, 52.1±10.9 years) from 57 families were selected for sustained hypertension (160.7 ± 22.9/99.5 ± 12.8 mmHg with 80% of patients under antihypertensive treatment), of early onset (40.7 ± 12.0 years), in the absence of obesity, diabetes mellitus, and secondary hypertension. Eight renin haplotypes were generated from three diallelic renin restriction fragment length polymorphisms (RFLPs) (TaqI, Hinfi, HindIII) located throughout the renin gene. The allelic concordance between the sib pairs was analyzed by identity by state relationships for 98 sib pairs (41 for 41 couples, 39 for 13 trios, 18 for 3 quartets). Allelic frequencies in the 57 hypertensive probands were similar to those observed among 102 hypertensive subjects studied previously. Six of eight possible haplotypes were observed, the informativity of the marker corresponded to 70% of heterozygosity. Allelic concordance for all sib pairs according to sibship size was not significantly different from that expected under the hypothesis of no linkage (t = 0.52, P = 0.15) reflecting only a small excess of renin alleles shared by the hypertensive sibs (1.44 ± 0.6 vs 1.36 ± 0.6). Likewise the linkage hypothesis was unsupported by weighted estimates to correct for possible bias due to large sibship size. Thus, the sib pair analysis suggests that the renin gene does not have a frequent role in the pathogenesis of essential hypertension; further more powerful linkage studies or other approaches will be needed to detect contributions at the renin locus to the heritability of essential hypertension.     

Studies on NK cell purification by negative selection in human peripheral blood

We have attempted to improve negative selection procedures for the large scale purification of human CD _in3 ^– CD56⁺ NK cells. In a series of experiments, purifications of NK cells from 10⁸ PBMC were performed by T cell depletion using either direct or indirect anti-CD3 labeling and the Magnetic Activated Cell Separation (MACS) procedure. Contaminating CD3⁺ cells were still present using either one of these two different T cell depletion protocols as shown by phenotyping IL-2 supplemented cell cultures on day 12. A second cycle of purification was therefore added. When MACS and Dynabeads were compared as complementary procedures to the first MACS cycle starting with 10⁸ cells, the Dynabeads method was found to be superior to the MACS with regard to the elimination of residual T cells. Starting from 10⁹ PBMC, we showed that this MACS+Dynabeads procedure gave similar satisfactory results when compared to the scaling-up of a previously established two steps procedure using Dynabeads. These two approaches (MACS+Dynabeads and 2 cycles of Dynabeads) have been also tested in a clinical setting to purify NK cells from cancer patients prior toin vitro expansion. The results indicate that the two methods are equivalent with respect to purity and recovery rate; a slight advantage in terms of feasibility was found in favor of 2 cycles of Dynabeads.     

Production of ψ-linolenic acid by the fungus Mucor rouxii on cheap nitrogen and carbon sources

Summary The production of -linolenic acid (GLA) by the fungus Mucor rouxii CBS 416.77 was studied on low budget nitrogen and carbon sources, i.e. rape meal, cocos expeller and two types of yeast extract (nitrogen sources), and starch, starch hydrolysate, beet molasses and cocos expeller (carbon sources). As references, Difco yeast extract and glucose were used. In flask cultivations the three yeast extracts were fully interchangeable, while the Difco yeast extract (the most expensive of those tested) gave a higher productivity of GLA in fermentor cultures (14 mg·l^–1·h^–1). The yield of lipids and GLA were increased in the order yeast extract < rape meal < cocos expeller. Thus the amount of lipid increased from 0.56 to 2.8 g·l^–1, and that of GLA from 0.15 to 0.33 g·l^–1. Use of beet molasses or cocos expeller as carbon sources gave poor growth. Starch and starch hydrolysate resulted in better productivity of GLA than glucose (4.7 and 4.9 compared to 3.4 mg·l^–1·h^–1). Offsprint requests to: A.-M. Lindberg     

Antisera Against the Indolealkylamines: Tryptophan, 5-Hydroxytryptophan, 5-Hydroxytryptamine, 5-Methoxytryptophan, and 5-Methoxytryptamine Tested by an Enzyme-Linked Immunosorbent Assay Method

Antisera were raised against tryptophan, 5-hydroxytryptophan, 5-hydroxytryptamine, 5-methoxytryptophan, and 5-methoxytryptamine, by conjugating each molecule to bovine serum albumin and to human serum albumin via glutaraldehyde, in such a way as to preserve the original part. Antibody specificity was tested with the enzyme-linked immunosorbent assay method. The specificity of each anti-indolealkylamine-glutaraldehyde antibody was established with competition experiments by using an adsorbed immunogenic conjugate and indolealkylamines either free or conjugated with poly-L-lysine. The nonconjugated compounds were poorly recognized. In the same way, the nonreduced conjugates always appeared less immunoreactive than the reduced ones. Calculated from the specificity study of each antiserum, the cross-reactivity ratios were found to be smallest for the most immunoreactive conjugates. Thus, a specific immune response was defined for each compound belonging to the same metabolic pathway.