Relative Humidity and the Killing of Bacteria: The Survival of Damp Serratia marcescens in Air

The viability of washed moist cells of Serratia marcescens after storage has been measured in relation to variations in the prior treatment of the cells and in conditions of storage. The factors considered were: (i) water content during storage; (ii) method of arriving at water content (partial drying in vacuum or freeze-drying and addition of water); (iii) presence or absence of air during storage.

Increasingly rapid decay occurs as the water content at which the cells are stored is diminished from above 90% to 20 or 30% (“critical” water content). It occurs in presence or absence of air and it occurs whether the final water content is approached by removal of water from wet cells or by addition of water to freeze-dried cells.

The rate of decay during storage at 20 to 30% water is somewhat diminished by the presence of air (“protective” effect of air).

As the water content is further reduced to less than 10%, the stability of cells stored in a vacuum approaches that of wet cells. In presence of air the reverse is true: the stability decreases until at less than 1% water, the decay rate is about as great as at the “critical” water content (“toxic” effect of air).

Particularly rapid decay of S. marcescens at the “critical” water content has escaped attention in aerosol studies because accurate control of relative humidity (RH) in this region, RH 94 to 99%, is virtually impossible in such studies. On the other hand, values of decay rates referred to measured water contents are quite unreliable in the 20 to 80% RH zone because the corresponding variation of water content is too small to measure reliably. Thus data of the kind reported in this paper cannot be directly compared to the published results of studies of air-borne bacteria, although they are relevant to the practical question of air-borne infection in humid atmospheres.

     

ELECTRON MICROSCOPE STUDY OF MITOSIS IN SEA URCHIN BLASTOMERES

The fine structure of cells at different stages of the mitotic cycle was studied in the blastomeres of 6-hour-old embryos of the sea urchin Strongylocentrotus purpuratus. The material was fixed in 1 per cent osmium tetroxide in sea water, buffered with veronal-acetate to pH 7.5, embedded in Araldite, and sectioned with glass knives. The aster, as it forms around the centriole, has the appearance of the endoplastic reticulum, with elements oriented radially from the centrosphere to the periphery of the cell. Anaphase structures described include the kinetochores, with bundles of fine filaments extending toward the centrioles, as well as continuous filaments passing between the chromosomes. Two cylindrical centrioles composed of parallel rods are present in each of the anaphase asters. At late anaphase, elements of the endoplasmic reticulum condense on the surface of the chromosomes to form a double membrane which already at this stage possesses pores or annuli. At telophase bundles of continuous filaments can be seen in the interzonal region. These filaments, as well as those associated with the chromosomes, have a diameter of approximately 15 mµ, and appear physically different from the astral structure.     

Improved Temperature-Gradient Incubator and the Maximal Growth Temperature and Heat Resistance of Salmonella

An improved all-metal temperature-gradient incubator produces its gradient by means of a bar permanently installed in a near-vertical position with its lower end in a cool constant-temperature water bath and with thermostatically controlled heaters near its top. Bolts hold the incubator in contact with the temperature-gradient bar, and polyurethane foam insulates the entire assemblage during use. Maximal growth temperatures of 34 representative strains of Salmonella were found to be between 43.2 and 46.2 C. In an agar medium with an initial level of 10⁶ cells per milliliter, no strain survived 50 C for 48 hr. S. senftenberg 775W showed no greater heat resistance at or near 48 C than did other species or other S. senftenberg strains. However, it was considerably more resistant than other strains at 55 C.     

CALAMOPHYTON IN THE MIDDLE DEVONIAN OF NEW YORK STATE

Specimens of Calamophyton from the Middle Devonian Ashokan Sandstone near Kingston, Ulster County, N.Y., are shown to belong to C. bicephalum Leclercq and Andrews, a Belgian species. Steel needles and a stereoscopic binocular microscope were used to follow the path of the forking leaves and branching sporangiophores of the specimens through the matrix. The terete leaves dichotomized two to three times in more than one plane. Sporangiophores dichotomized once. Each branch bore three recurved lateral branches which in turn bore two sporangia. Each branch terminated in an elongate, filiform projection. Sporangia apparently dehisced longitudinally. Their walls were composed of elongate cells. Their spherical spores ranged from 86 to 166 μ in diameter and bore a trilete mark. Ornamentation consisted of coni and spinae up to 4.5 μ long. They resembled dispersed spores of Dibolisporites gibberosus var. major Richardson. This is the second occurrence of Calamophyton bicephalum and the first account of its spores. It is the second report of the genus in North America.     

Lipids of isolated neurons

1. Lipids were extracted from neurons isolated from the lateral vestibular nucleus of ox (Bos taurus L.) and the ganglia of Aplysia punctata Cuvier. 2. Thin-layer chromatography of ox-neuron lipid revealed three major fractions corresponding to neutral lipid, phosphatidylethanolamine and phosphatidylserine. Part of the phosphatidylethanolamine was present as the plasmalogen. 3. Aplysia-neuron lipid contained neutral lipid, phosphatidylethanolamine and phosphatidylserine. Both phospholipids appeared to be present predominantly as the plasmalogen form. 4. The fatty acids of alkali-labile lipids of ox neurons were examined by gas–liquid chromatography. The major fatty acids were oleic acid, stearic acid and palmitic acid.     

The activities of nicotinamide mononucleotied adenylyltransferase and of nicotinamide–adenine dinucleotide kinase in the livers of rats subjected to different hormonal treatments

1. The activities of NMN adenylyltransferase and of NAD(+) kinase have been measured in the livers of adrenalectomized or alloxan-diabetic rats and in the livers of rats treated with glucagon, pituitary growth hormone or thyroxine. 2. The activities of these enzymes have been compared with the effects of the same treatments on the nicotinamide nucleotide concentrations in the liver. 3. Alloxandiabetes (+37%) and thyroxine (+27%) both increased the activity of NMN adenylyltransferase. The other treatments were without effect on this enzyme. 4. Only thyroxine increased the activity of NAD(+) kinase significantly (+26%) although both adrenalectomy and glucagon tended to increase its activity. 5. The activity of NAD(+) glycohydrolase was measured in the livers of diabetic rats, and in the livers of rats treated with either growth hormone or thyroxine. Of these treatments, only growth hormone altered the enzyme activity (+26%, calculated on a total hepatic activity basis). 6. Female rats had a greater hepatic NAD(+)-kinase activity than males but there was no sex difference with respect to NMN adenylyltransferase. 7. The lack of correlation between the maximum potential activity of these three enzymes and the known changes of the nicotinamide nucleotides in each of the hormone conditions is discussed.     

Segregation products of male mice doubly heterozygous for the RB(6.16) and RB(16.17) translocations: Influence of sperm karyotype on fertilizing competence under varying mating frequencies

The meiotic segregants of male mice heterozygous for Rb(6.16)24Lub and Rb(16.17)7Bnr were viewed, for the first time, at first cleavage metaphase. Chromosomes were analyzed after G-banding, C-banding, and karyotyping. To study sperm aging effects, chromosomes of 202 one-cell zygotes derived from males mating at intervals of approximately 3,14, and 21 days were examined. At least 89.6% of sperm-derived complements were products of 2:2 segregation; at most, a possible 6.4% were 3:1 segregants. The six expected types of 2:2 segregants, both balanced and unbalanced, were equifrequent in the total zygote population derived from sperm of all ages. When the data were analyzed according to mating frequency, the 3-day sperm population considered most likely to be fresh showed a deficiency of the segregant nullisomic for chromosome 6 and disomic for chromosome 17, when compared with the reciprocal segregant (P < 0.025) as well as to all other 2:2 segregants (P < 0.05). However, these sperm fertilized in greater numbers (P < 0.01) than their reciprocal segregant (disomic for 6 and nullisomic for 17) in the 14-day sperm population. While sperm with chromosomal abnormalities are capable of fertilization, the competence of segregants nullisomic for 6 and disomic for 17 apparently depends on the prior storage period in the male. Further, the results suggest that the effect of aneuploidy on sperm function is dependent on the specific chromosome(s) involved.     

Stainless steel mesh supports high density cell growth and production of recombinant Mullerian Inhibiting Substances

Summary Stainless steel mesh supported the high density growth of anchorage dependent CHO fibroblasts without the use of a special culture system. CHO cells, designated B-9, containing an amplified genomic construct of the human gene for Mullerian Inhibiting Substance (MIS), grew to a high confluent density on stainless steel meshwork while producing substantial amounts of human recombinant MIS over a long period of time. The mesh could be easily coated with various extracellular matrix proteins, such as Laminin, Fibronectin, Collagen or Matrigel, which permitted the testing of the effects of surface modifications on cell yield and recombinant protein production. Since the amount of medium per surface area required for optimal cell growth is lower than for some large volume cell culture methods, media costs can be reduced using mesh. In addition, no special cell culture equipment or complex manipulations are required. Thus, the use of meshwork for anchorage-dependent cells can increase the efficiency of growth and decrease the cost of recombinant protein production. This work is supported by NIH grant CA 17393 and American Cancer Society grant PDT 221A to P. K. D. and NIH grant EY 06535 to J. E. Editor's Statement This approach to large scale, high density cultivation of cells, one of several which are based on increasing surface area of the cultures, allows the production of large amounts of recombinant product within a research laboratory with modest bulk culture capability.