Immunocytochemical and ultrastructural characterization of endocrine cells in chicken proventriculus

Summary The endocrine cells of the chicken proventriculus were investigated immunocytochemically, using the peroxidase-antiperoxidase technique on paraffin and semithin sections for light microscopy, and immunogold staining in osmium-fixed material for electron microscopy. The fixation procedure also allowed a detailed ultrastructural investigation. Twenty-three antisera were tested and 7 immunoreactive cell-types were identified: D-cells containing somatostatin-like peptide; EG-cells immunoreactive to anti-glucagon, anti-GLP1 and antineurotensin; NT-cells labelled only with anti-neurotensin; BN-cells containing bombesin-like material; ENK-cells showing met-enkephalin immunoreactivity; EC-cells reactive to anti-serotonin; and APP-cells positive to anti-avian pancreatic polypeptide. In addition, enterochromaffin-like (ECL) cells, were also detected by electron microscopy. The presence of ENK-cells and the ultrastructure of these and NT-cells are described for the first time in chicken proventriculus, and glucagon, GLP1 and neurotensin are shown to be colocalized in the EG-cells.     

Wide distribution of a 36-megadalton plasmid among clinical and environmental Spanish isolates ofLegionella pneumophila serogroup 1

With the eventual goal of characterizingLegionella pneumophila serogroup 1 plasmids at the molecular level, we have analyzed the plasmid contents of 78 clinical and environmental Spanish isolates. After selection of a suitable alkaline lysis method, we detected plasmids with approximate molecular weights of 25, 36, 40, 61, 80, 85, 90, and 95 megadalton (MDal). Several factors (i.e., wide temporal and geographic distribution, high frequency in both clinical and environmental isolates, and apparent high copy number after subculturing) make the 36 MDal type IA plasmid an appropriate plasmid for further molecular studies.     

A new family of repetitive nucleotide sequences is restricted to the genus Zea

We have isolated a new family of moderately repetitive nucleotide sequences (about 2500 copies per haploid genome) specific to the genus Zea and absent in other graminaceous species. These sequences are interspersed in the genome and they show the same genomic organization pattern and similar copy number in all the Zea species examined. These two facts, consistency in the copy number and the same organization pattern, would indicate on the one hand that these sequences were amplified before the divergence of Zea species, and on the other hand that maize and all the teosintes could be considered as the same evolutionary population. Independent clones corresponding to the repetitive sequences have been isolated and sequenced from a genomic library of the teosinte, Zea diploperennis. The repeats, flanked by HaeIII sites, are more than 70% G + C-rich, on average 253 bp long and show 78% similarity to each other. These repetitive sequences are in a highly methylated-C context and they present some features resembling those of coding sequences, such as high CpG and low TpA content, and similar codon usage to maize genes in one of the reading frames. Moreover, the repetitive probe hybridizes with RNA extracted from different tissues of maize and from teosinte, indicating that these repeats or similar ones are present in transcribed sequences.     

The R-banding pattern of the Chinese hamster Don cell line

Chinese hamster cells (Don line) were treated in vivo with 5-BrdU and 33258-Hoechst fluorochrome for obtaining the partial inhibition of condensation that causes the R-banding pattern. Untreated chromosomes were stained by a standard G-banding method. Statistical measurements show significant differences in the band numbers between the two treatments. The Don cell line in the authors' laboratory presents some karyotypical differences from Don cell lines studied by other authors.     

Elutriation of human keratinocytes and melanocytes fromin vitro cultured epithelium

Summary Human epidermal keratinocytes grown in culture and at different stages of differentiation are shown to be viably separated by elutriation. A specific fraction enriched in melanocytes was obtained. Elutriation of cells obtained fromin vitro cultured epithlium could prove useful in studies concerning the biochemistry and molecular markers of cells isolated from normal epithelium and from different pathologies.     

Effect of some antineoplastics on metabolic heme pathway

1. The porphyrinogenic ability of several antineoplastics used in the therapy of the different cancers was evaluated. The action of cyclophosphamide, busulfan and 5-fluorouracil on the amount and nature of the accumulated hepatic porphyrins and on the activity of delta-aminolaevulinate synthase (ALA-S), were estimated at different doses and times of drug treatment in 17-day-old chick embryos. 2. It was observed that cyclophosphamide produces a significant increase in the accumulation of hepatic porphyrins at different doses as well as in the activity of the ALA-S, at all the incubation times. Cyclophosphamide alters the pattern of porphyrins accumulated in the liver, where a coproporphyrin: protoporphyrin ratio higher than in the controls can be observed. 3. Busulfan increased the hepatic porphyrins accumulated in the liver but to a lesser degree than cyclophosphamide. 4. 5-Fluorouracil did not modify the hepatic porphyrin content when it was administered at doses up to 40 mg/embryo. 5. When the embryos were injected with busulfan or 5-fluorouracil no significant differences were observed in the activity of ALA-S up to 11 hr of incubation. 6. These results indicate that cyclophosphamide has a remarkable porphyrinogenic capacity in chick embryo while busulfan, notwithstanding the fact that it alters the haem pathway, it does so to a degree that does not impair the regulation of ALA-S activity. Fluorouracil seems to be non porphyrinogenic in this system, up to 40 mg/embryo.     

Comparative studies of selective media for recovery of Staphylococcus aureus from natural waters

Several selective media currently used for the enumeration of Staphylococcus aureus from different sources were evaluated in order to establish their quantitative recovery, specificity and degree of selectivity, using different types of water samples. The highest selectivity and reliability in the enumeration of Staph. aureus from the samples was obtained on Borrego-Florido-Romero-0 (BFR-0) and KRANEP agars. The method that produced the highest recovery of Staph. aureus was BFR-0 agar with membrane filter and incubation at 36 degrees C for 48-72 h.     

Gene activity of polytene chromosomes in Drosophila species of the obscura group

The polytene chromosome puffing patterns of Drosophila guanche were established and compared with those of Drosophila subobscura. A total of 150 loci, active in some of the 17 developmental stages studied, were described and 23 of them were found to form the characteristic puffing pattern of D. guanche. Taking into account the number of puffs as well as the gene activity of each chromosome and the total gene activity, D. guanche seems to be less active than D. subobscura. Although both species show a degree of homology in their puffing patterns lower than that found for sibling species, the degree of homology is stronger than that between species belonging to the same group but to different subgroups. Thus, D. guanche and D. subobscura must be considered as phylogenetically closely related species, belonging to the same subgroup.     

Polyamine Metabolism and Osmotic Stress : II. Improvement of Oat Protoplasts by an Inhibitor of Arginine Decarboxylase

We have attempted to improve the viability of cereal mesophyll protoplasts by pretreatment of leaves with dl-α-difluoromethylarginine (DFMA), a specific `suicide' inhibitor of the enzyme (arginine decarboxylase) responsible for their osmotically induced putrescine accumulation. Leaf pretreatment with DFMA before a 6 hour osmotic shock caused a 45% decrease of putrescine and a 2-fold increase of spermine titer. After 136 hours of osmotic stress, putrescine titer in DFMA-pretreated leaves increased by only 50%, but spermidine and spermine titers increased dramatically by 3.2- and 6-fold, respectively. These increases in higher polyamines could account for the reduced chlorophyll loss and enhanced ability of pretreated leaves to incorporate tritiated thymidine, uridine, and leucine into macromolecules. Pretreatment with DFMA significantly improved the overall viability of the protoplasts isolated from these leaves. The results support the view that the osmotically induced rise in putrescine and blockage of its conversion to higher polyamines may contribute to the lack of sustained cell division in cereal mesophyll protoplasts, although other undefined factors must also play a major role.