pIN32: a cointegrate plasmid with IncHI2 and IncFII components

An Enterobacter cloacae strain isolated from the faeces of a child with diarrhoea in Indonesia contained a transferable 216 MDa plasmid, pIN32, exhibiting IncHI2 phenotypic characters, including temperature sensitivity of transfer and the expression of H serotype pili at a repressed level. A derivative plasmid (pIN32-1), which had lost the IncHI2 phenotype, and contained only 60 MDa of the original replicon, was obtained after mating at 37 degrees C. It was IncFII, showed regions of homology with plasmid R100, determined IncFII serotype conjugative pili constitutively and was transfer-derepressed. After overnight growth at 37 degrees C in non-selective medium, pIN32 gave rise to another derivative, pIN32-2 (size 184.3 MDa), which retained the IncHI2 phenotype and several other pIN32 characters.     

Species-Specific Restriction Site Polymorphism in Root-knot Nematode Mitochondrial DNA

Research was initiated to physically characterize the mitochondrial genomes of several Meloidogyne spp. and host-races, to address questions regarding their systematics and dispersal, and to assess the possibility of developing molecular diagnostics for these nematodes. Techniques were developed for purification and rapid detection of mitochondrial DNA from root-knot nematodes. Mitochondrial DNAs among Meloidogyne spp. were demonstrated to exhibit extensive divergence. The potential for using the rapidly diverging mitochondrial genomes as a diagnostic assay for M. incognita, M. hapla, M. arenaria, and M. javanica is discussed.     

A comparison between the hot and cold water soluble fractions of two locust bean gum samples

Locust bean gum extracted from two carob flours from eastern and western Mediterranean sources were fractionated on the basis of their solubility in water. Weight-average molecular weights determined by sedimentation equilibrium were about 300 000 for both the hot water and cold water soluble fractions, whereas a commercial sample of guar gum had a molecular weight of 700 000. Their values were lower than would be predicted from Mark-Houwink relationships where molecular weights were originally determined by light scattering.

The hot water soluble fraction from the eastern Mediterranean flour showed unexpected rheological behaviour. It had an extremely high Huggins' constant and a different relationship between the coil overlap parameter and the zero shear rate viscosity compared with previously reported results for galactomannans. Both effects may be explained by the anomalously low intrinsic viscosity of this fraction when determined by a Huggins' extrapolation. The use of the Kraemer extrapolation gave significantly higher intrinsic viscosities for this particular sample. Gels formed from the two hot water soluble fractions with κ-carrageenan had similar rheological properties.     

Changes in the morphology and synthetic activity of cultured rat tail tendon

Summary Isolated single fascicles from tail tendons of young rats were freed of epitenon cells and cultured in vitro for up to 7 days. The tissue remained viable, as judged by the structural integrity of cell organelles and the ability to synthesize DNA and glycosaminoglycans (GAG). The rate of DNA synthesis peaked after 2 days in culture and decreased slowly thereafter. Concomitantly, an increase in cell number was noted at the periphery of the fascicle. GAG production also increased during culture, sulphated GAG being increased proportionately more than hyaluronic acid. Dermatan sulphate was the predominant sulphated GAG in freshly isolated fascicles, but in cultured tissue, the newly synthesized sulphated GAG was more sensitive to degradation by chondroitinase AC and had an increased electrophoretic mobility. Fine structural changes were observed in cultured tissues such as the retraction of cell processes. rounding up of cell bodies and the appearance of gaps between collagen fibrils. Cultured tenocytes also frequently contained apparently phagocytized collagen fibrils which were not seen in freshly isolated fascicles, and this appearance was suggestive of collagen degradation occurring in vitro, although no change in the total hydroxyproline content was noted. The data show that when individual fascicles are cultured in vitro they undergo a process of matrix remodelling which has features in common with events occurring in vivo when tendons have been surgically manipulated.     

Isolation and characterization of an α-satellite repeated sequence from human chromosome 22

We constructed a library in IL47.1 with DNA isolated from flow-sorted human chromosome 22. Over 50% of the recombinants contained the same highly repetitive sequence. When this sequence was used to probe Southern blots of EcoRI-digested genomic DNA, a ladder of bands with increments of about 170 bp was observed. This sequence comigrates with satellite III in Ag⁺/Cs₂SO₄ gradients and may account for at least part of the 170 bp Hae III ladder seen in isolated satellite III DNA. Partial sequence analysis revealed homology to the 171 bp monomeric repeat unit of -R1-DNA and the X specific -satellite consensus sequence. After low stringency in situ hybridization, silver grains were found over the centromeres of a number of chromosomes. Under high stringency conditions, however, the labeling was concentrated over the centromeric region of chromosome 22. This localization was confirmed using DNA from a panel of human/hamster cell lines which showed that the homologous 2.1 and 2.8 kb EcoR1 restriction fragments were chromosome 22 specific. These clones therefore contain chromosome 22 derived -satellite sequences analogous to other chromosome-specific satellite sequences described previously.     

Lectin-binding activity and antibody response with Pseudomonas paucimobilis and Flavobacterium multivorum

Pseudomonas paucimobilis and Flavobacterium multivorum (formerly Group IIK biotype 1 and biotype 2, respectively) showed lectin-binding activity with Helix aspersa and no activity with eight other well-characterized lectins. Microagglutination titrations and immunodiffusion precipitation revealed specific antibody activity to immunizing antigens. However, no major antigens were found to be common from crude antigen preparations of P. paucimobilis and F. multivorum when tested against opposing antisera.     

Developmental Stability of DROSOPHILA MELANOGASTER under Artificial and Natural Selection in Constant and Fluctuating Environments

Populations of Drosophila melanogaster in constant 25° and fluctuating 20/29° environments showed increases in developmental stability, indicated by decreases in bilateral asymmetry of sterno-pleural chaeta number. In both environments, rates of decrease in asymmetry were greater under natural selection (control lines) than under artificial stabilizing selection. Overall mean asymmetry was greater in the fluctuating environment.—There was no evidence that decreased asymmetry was due to heterozygosity, and the decline in asymmetry was not explained by the decline in chaeta number in the lines under only natural selection. However, the decline was consistent with changes in total phenotypic variance and environmental variance.—The divergence between lines after 39 generations of selection was seen in differences in asymmetry and also in the genotype-environment interaction expressed in cross-culturing experiments.     

Determination of pili by conjugative bacterial drug resistance plasmids of incompatibility groups B, C, H, J, K, M, V, and X.

Representative plasmids from incompability groups B, C, H, J, K, M, V, and X were transferred to "bald" strains of Escherichia coli or Salmonella typhimurium. By using a new technique, pili were detected by electron microscopy for each incompatibility group. Morphology varied but was similar for plasmids within a group. These findings suggest that all conjugative plasmids in the Enterobacteriaceae may determine pili.     

Stimulatory modulator-requiring cyclic nucleotide-independent protein kinase: Partial purification from fetal calf hearts and comparison with various protein kinases

A stimulatory modulator-requiring cyclic nucleotide-independent protein kinase was purified over 400-fold from the extract of fetal calf hearts by the steps of DEAE-cellulose and Sephadex G-100 chromatographies. The enzyme was activated by stimulatory modulator of cGMP-dependent protein kinase. Inhibitory modulator (protein inhibitor) of cAMP-dependent protein kinase, calcium, phosphatidyl serine and cyclic nucleotides were without effect. The enzyme (3.2 S) was much smaller than the holoenzymes of cGMP- and cAMP-dependent protein kinases. This new species of enzyme thus appears to be similar to the putative catalytic subunit of cGMP-dependent protein kinase previously reported.