Progression of Epiphytic Microflora in Wheat and Alfalfa Silages as Observed by Scanning Electron Microscopy

Wheat and alfalfa silages were examined by scanning electron microscopy and standard methods of microbial enumeration. Epiphytic microflora were present at levels of 10⁶ to 10⁸/g in the fresh-cut plants. This flora was initially observed microscopically primarily on the surfaces. After 4 days of fermentation, lactic acid bacteria were observed on the surface in high concentrations near open stomata and throughout the interior mesophyll air sac spaces. At 4 days, populations on interior surfaces were restricted to the exterior surfaces of the air sacs. After 8 days the mesophyllic cells showed marked deterioration, and bacteria were observed on their inner surfaces. At 32 days, the end of the fermentation, vascular bundles and epidermal cells remained intact whereas stomata and mesophyllic cells were collapsed and often contained microorganisms. It is concluded that the interior of the leaves offers substantial nutritional and environmental advantages to epiphytic flora and is an important if not major deterioration site in fermented products. Since little deterioration of exterior surfaces was observed, these sites may play a minor role in supplying nutrients for microbial growth.     

Evidence for Cometabolism in Sewage

A procedure was developed to demonstrate cometabolism in models of natural ecosystems. The procedure involves showing the formation of metabolic products in high yield and the lack of incorporation of substrate carbon into cellular constituents. Samples of four ¹⁴C-labeled herbicides (trifluralin, profluralin, fluchloralin, and nitrofen) were incubated with sewage aerobically and under discontinuous anaerobiosis for 88 days, and fresh sewage was added at intervals. Products were formed from each of the herbicides in nonsterile, but not in sterile, sewage. The yield of recovered products reached 87% for profluralin and more than 90% for fluchloralin and trifluralin, and the number of products ranged from 6 for nitrofen to 12 for fluchloralin. Concentrating the sewage microflora 40-fold greatly enhanced the rate of conversion. None of the radioactivity from the herbicide entered the nucleoside pool of the sewage microflora. The lack of incorporation of substrate carbon into cells and the almost stoichiometric conversion of the substrate to organic products indicate that members of the microbial community were cometabolizing the test compounds.     

Regulation of self-recognition by T-cell hybrids

Somatic cell hybrids were prepared between BW 5147, an AKR T lymphoma, and purified T cells from three sources: spleen cells exposed to sheep red blood cells, lymph node cells from mice sensitized to ovalbumin, and spleen cells of mice injected with azobenzenearsonate-IgG. Hybrid lines expressed constitutive markers of both parents which include H-2 antigens and the isoenzymes glucose phosphate isomerase and isocitrate dehydrogenase. Furthermore, they expressed both parental alleles of Thy 1, a differentiation antigen. Many of the hybrid lines formed rosettes with mouse erythrocytes. T-cell hybrids did not bind human or chicken red blood cells, though they did rosette with sheep erythrocytes to the same extent as with mouse red cells. We interpret the latter reaction as due to recognition of shared antigens by the murine T cells. This form of self-recognition is influenced by culture conditions and is expressed optimally by cells in late logarithmic phase of growth.     

Specificity of Insertion by the Translocatable Tetracycline-Resistance Element Tn10

Genetic analysis of 131 independent transpositions of the tetracycline-resistance element Tn10 from a single site in phage P22 into the histidine operon of Salmonella typhimurium reveals that Tn10 insertions are not randomly distributed along this chromosomal target. The insertions occur in 22 different "clusters"; insertions within each cluster are very tightly linked in recombination tests. Tn10 insertions are not evenly distributed among the identified clusters. The existence of these clusters suggests that this chromosomal target contains particular genetic signals that guide Tn10 to particular preferred positions for insertion. Insertions within each cluster occur in both orientations with roughly equal frequency.--The relationship among different insertions within each cluster has been examined. The resolution of genetic mapping places an upper limit of about 50 basepairs on the distance between different insertions within a cluster. Different insertions within a cluster usually have the same reversion frequency; however, heterogeneity in reversion frequency has been detected in at least two clusters. For most clusters, the available data are consistent with the simple possibility that all insertions within a cluster are at identical positions; however, the data do not exclude other possibilities.     

Metabolic and temporal studies on pancreatic exocrine cells in culture

Summary An approach is described whereby cells with definitive markers are followed from their source through dissociation and fractionation, then during long-term maintenance in vitro. Such sequential studies should enable investigators to define factors regulating proliferation and function of specific cells since ambiguity concerning identity is readily avoided.Pancreatic cells of guinea pigs were isolated by enzymic dissociation, and exocrine cells were enriched by centrifugation with solutions of serum albumin. Resulting populations consisting of up to 95% exocrine cells were then incubated with gyration to produce aggregates, and these were seeded to standard culture plates for further study. Colonial aggregates of exocrine epithelia develop in culture and can be maintained for 20–30 days. The cells exhibit changes with time that are qualitatively similar to those known to occur during serial cultivation of diploid fibroblastlike cells from human and other species. The uptake of tritiated thymidine decreases with maintenance time. Autoradiographic examination indicates that this is due to a reduction in the number of epithelial cells incorporating the isotope. Cell diameters increase from an average of 21 m at day 0 to 44 m by day 26, and a marked increase in heterogeneity of this parameter is also evident. Cellular DNA and protein accumulate during the same interval. Incorporation of tritiated leucine during 24-h exposures increases until about the 10th day in vitro and remains relatively constant for at least 2 weeks thereafter.The data are consistent with the hypothesis that exocrine pancreatic cells like other diploid cells in culture, progress to terminal differentiation under the culture conditions employed. The role of physical, nutritional, and humoral evironmental factors on this process will be the subject of future reports.Supported in part by National Cancer Institute Contracts NO1-CP-43231 and NO1-CP-65751     

Tissue microenvironment and the genetic control of hair pigment patterns in mice

This study was conducted to assess microenvironmental variability within integumental tissue of genetically identical mice with respect to a specific cellular response: cyclic synthesis of yellow and black pigment by hair bulb melanocytes. Crosses were performed within and between inbred strains of mice that were isogenic with the exception of a single gene substitution at the agouti locus. Agouti locus genes included the A^vy, A^w, A, a^td, a^t, a^x, a^m, and a alleles. The pigment patterns of dorsal, flank, and ventral hairs of the first and third hair generations and of hairs growing in special integumentary areas such as the pinna, tail, and hind foot were studied. It was found that the amount of yellow pigment synthesized by hair bulb melanocytes within genetically identical mice is both agedependent and conditioned by the integumentary environment. Furthermore, the special integumentary regions produce hairs with a variety of pigment patterns in which the distribution and relative amounts of black and yellow pigments do not necessarily conform to dominance relationships expected among agouti locus alleles as judged by their effects on the pigmentation of the dorsal pelage. We conclude that within genetically uniform integumental tissues, microenvironmental differences occur and are reflected as alterations in the metabolic pattern of differentiated cells.     

Specific protection from phenocopy induction by heat shock

Mild heat treatments applied to whole animals or cell cultures of Drosophila prior to lethal heat shocks result in increased survival and protection against phenocopy induction. The optimal condition for the preliminary mild heat treatment is that which induces the synthesis of heat-shock proteins but does not turn off the protein synthesis that is in progress. Recovery of protein synthesis but not RNA synthesis following a drastic heat shock is much enhanced by the pretreatments. The results suggest that the protection for survival and against phenocopy induction is due to storage of messenger RNA.     

Terminal Redundancy Heterozygotes Involving the First-Step-Transfer Region of the Bacteriophage T5 Chromosome

Individual progeny of two-factor crosses between A1am and A2am T5 phages give rise to bursts containing more than one type of plaque. The simplest explanation for these mixed bursts is that the A1 and A2 genes are located within the terminally repeated portion of the T5 genome and that the mixed bursts are made by "terminal redundancy heterozygotes". The observation of genetic heterozygosity means that the A1 and A2 genes are repeated intact. This implies that the terminal segments of T5 are genetically interchangeable.     

Xanthine dehydrogenase accumulation in developing Drosophila eyes

Xanthine dehydrogenase activity is assayed by following the oxidation of pterin to isoxanthopterin by spectrofluorometry at the reaction's wavelength peaks: excitation, 344 nm; emission, 412 nm. The method is sensitive to less than 0·1 μU of activity (0·1 pmol/min) and allows the assay of Drosophila imaginal disk homogenates.While the larval eye disk contains less than 0·1 per cent of the individual's XDH, the developing eye becomes a major store, with 30 per cent of the individual's activity by the time of eye pigmentation. The data suggest a basis for the well-known non-autonomous action of the gene rosy, the structural gene for XDH: the enzyme is synthesized in an organ of primary gene expression, and transported through the haemolymph to the eye of the pupa and pharate adult.     

Peptidoglycan compositions of a new strain of Arthrobacter crystallopoietes during sphere-rod morphogenesis

Arthrobacter crystallopoieties ATCC 15481 was used to isolate a new strain, designated Arthrobacter crystallopoieties EPSR-16, which had a mass doubling time in brain heart infusion broth and in glucose/salts/yeasts extract medium of 30 min compared to 2.40 h for the parent strain in similar media. The growth rates for the new strain and for the parent were close to 12 h in glucose/salts medium. The new strain formed well-separated cocci and diplococci in glucose/salts medium, and upon nutrient shift-up all the cells in the population gradually changed into well-separated rods of regular shape. In the spherical state the cell wall peptidoglycan of the new strain contained lysine and no diaminopimelic acid. A gradual loss in lysine and a gain in diaminopimelic acid occurred during morphogenesis. Diaminopimelic acid became predominant in the cell wall during balanced growth in the rod state.