Joint transfer of genetic markers in Bacillus subtilis

Takahashi, I. (McMaster University, Hamilton, Ontario, Canada). Joint transfer of genetic markers in Bacillus subtilis. J. Bacteriol. 91:101-105. 1966.-To compare the processes of genetic incorporation in transduction and transformation in Bacillus subtilis, several groups of linked markers were selected and the degree of linkage was determined by the two means of genetic exchange. Bacteriophage PBS 1 was used in transduction experiments. In all cases, frequencies of joint transfer, as expressed by the cotransfer index or by percentage of joint transfer, were higher in transduction than in transformation. With a pair of closely linked markers, the frequency of joint transduction was only slightly higher than that of joint transformation. On the other hand, a considerably higher degree of linkage was obtained by transduction when loosely linked markers were examined. It appears that the size of donor chromosome transferred by transducing phage particles is much larger than that incorporated by recipient cells in transformation. It is suggested that transduction in B. subtilis may be a useful tool in extending further the linkage groups established by the transformation technique.     

Stimulating effects of insulin and low-density lipoprotein on cell growth and macromolecular syntheses of HeLa cells cultured in K(+)-depleted medium

The influence of the intracellular K+ concentration on the effects of growth factors (insulin, EGF, hydrocortisone, and transferrin) and LDL on growth of HeLa cells was investigated. Upon replacement of K+ in a chemically defined medium (K(+)-CDM) by Rb+ (Rb(+)-CDM), about 80% of the intracellular K+ was replaced by Rb+ within 24 h, but showed no further change in the next 24 h, irrespective of addition of dialyzed calf serum (5%) or growth factors to the medium. In Rb(+)-CDM, cell growth and DNA synthesis were greatly suppressed, although cell viability was not significantly altered for 72 h. The suppression of cell growth was partially restored by addition of serum, insulin (5 micrograms/ml), or LDL (2.5 mg/ml) to Rb(+)-CDM. A combination of serum and insulin or insulin and LDL stimulated cell growth to approximately the level in K(+)-CDM without any addition, but a combination of serum and LDL did not have more effect than that of serum alone. Unexpectedly, other factors were ineffective in stimulating growth in Rb(+)-CDM. In Rb(+)-CDM, the effect of insulin was lost in 24-48 h, whereas that of LDL persisted for at least 96 h. Insulin and LDL also enhanced growth in K(+)-CDM. After cessation of cell growth in Rb(+)-CDM for 24 h, addition of insulin and/or LDL markedly restored cell growth and DNA synthesis. Therefore, insulin and LDL may stimulate certain mechanisms required for cell growth that can operate in K(+)-deficient conditions.     

Freemartinism among singleton bovine females born from multiple embryo transfer

Heterosexual chimerism among singleton females produced by multiple nonsexed embryo transfer (MNET singleton females) was investigated using chromosome typing and PCR (polymerase chain reaction)-amplification of male-specific DNA (msDNA). Of the 22 animals tested, 21 were classified as normal by both methods (i.e., showing no male cells among 100 metaphase spreads in chromosome typing and being msDNA negative in PCR). No morphological abnormalities of the genital organs were observed among 19 MNET single females. One MNET singleton female was, however, classified as a freemartin by PCR (male-specific DNA positive), but it was classified as normal cytogenetically. This individual probably had a low degree of heterosexual chimerism, and it seems that the chimerism derived from MNET was difficult to diagnose by chromosome typing, although it was detectable by PCR. The genital organs of this individual (15-mo-old Aberdeen Angus) were normal in form (both external and internal) and size. However, a very small structure, resembling seminiferous tubule, was found in the left ovary. It may be concluded that most MNET singleton females are expected to have normal reproductive function.     

Localization of the antigen for the monoclonal antibody ②9F11-B-E4 interspecific cross-reaction in the freshwater triclads

The localization of the antigen for monoclonal antibody 9F11-B-E₄ was clarified by immuno-electron microscopy. The antigens were localized on the mitochondria and Golgi bodies in the male germ cells and on the secretory granules of various glands cells in the penis bulb and subepidermal parenchymal tissue of Phagocata vivida. The results of the interspecific cross-reaction tests with seven other freshwater triclads showed that these secretory granules are species-specific. A positive interspecific reaction was showed with Dugesia (family Dugesiidae), but not with Polycelis within the same family Planariidae which suggests the position of Phagocata within the Planariidae needs to be reassesed.     

D-Amino acid production from racemic amino acids by a microbial asymmetric degradation

To obtain D-amino acid from a mixture of racemic amino acids, Proteus vulgaris, having the ability of selective degradation of L-isomer in various DL-amino acids mixtures, was used as a biocatalyst. D-Isomer crystals of valine, leucine, isoleucine, histidine and ornithine were isolated at a good yield (25~41%) from the reaction mixture after 24~72 hours.     

Changes in cytokeratin expression in epidermal keratinocytes during wound healing

In order to investigate the re-epithelialization process during wound healing, the hair on the back of guinea pigs was shaved and then excisional wounds were made through the entire thickness of the skin. Histological changes were observed and changes in the expression of different cytokeratin polypeptides were examined using an immunohistochemical technique. Immunohisto chemical study revealed that the proliferating and migrating keratinocytes expressed the same cytokeratins as the basal cells of normal epidermis. In addition, the entire epidermis of fairly remote areas from the edges of the wound where no thickening was observed showed a temporarily abnormal staining pattern. The suprabasal cells in the regenerating epidermis temporarily expressed cytokeratins not only specific for suprabasal cells but also specific for basal cells. The cytokeratins expressed in normal basal keratinocytes were also present in the thickened granular layers. These data indicate that the expression of cytokeratins in the epidermal keratinocytes (even in fairly remote areas from the wound edges) changes during wound healing, that the origin of the migrating keratinocytes from the remaining epidermis seems to be the basal cells in the epidermis, and that the appearance of keratohyalin granules is not related to changes in cytokeratin expression.     

Varicella infection in a children's hospital: prevention by vaccine and an episode of airborne transmission

Varicella vaccine was used safely and effectively for preventing ward infection with varicella. Ward infection was experienced 34 times in 5 years between 1977 and 1982. During these ward infections, varicella developed in 4 of 142 patients who received vaccine, 21 of 47 patients who did not receive vaccine and 1 of 9 who received transfusion with vaccine-boostered blood. Of the 142 vaccinated patients, the four in whom varicella developed showed symptoms 3 to 10 days after vaccination, indicating that vaccination had been too late. Details of a ward infection with varicella by airborne transmission and its prevention by vaccination are presented.