ABSTRACT
Microbial communities found on the surface of overwintering plants may be exposed to low temperatures as well as multiple freeze-thaw events. To explore the adaptive mechanisms of these epiphytes, with the objective of identifying products for freeze-protection, enrichment libraries were made from frost-exposed leaves. Of 15 identified bacteria from 60 individual clones, approximately half had ice-association activities, with the great majority showing high freeze-thaw resistance. Isolates with ice nucleation activity and ice recrystallization inhibition activity were recovered. Of the latter, two (Erwinia billingiae J10, and Sphingobacterium kitahiroshimense Y2) showed culture and electron microscopic evidence of motility and/or biofilm production. Mass spectrometric characterization of the E. billingiae extracellular polymeric substance (EPS) identified the major proteins as 35 kDa outer membrane protein A and F, supporting its biofilm character. The addition of the EPS preparation increased the freeze-thaw survival of the more susceptible bacteria 1000-10000 times, and protection was at least partially dependent on the protein component.
Subject(s)
Bacterial Proteins/chemistry , Biofilms/drug effects , Erwinia/physiology , Microbial Consortia/physiology , Sphingobacterium/physiology , Adaptation, Physiological , Bacterial Proteins/metabolism , Bacterial Proteins/pharmacology , Biofilms/growth & development , Chrysanthemum/microbiology , Escherichia coli/drug effects , Escherichia coli/growth & development , Freezing , Ice , Microbial Viability/drug effects , Microscopy, Electron , Plant Leaves/microbiology , Pseudomonas syringae/drug effects , Pseudomonas syringae/growth & development , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization , SymbiosisABSTRACT
We have investigated the toxic effects of trichloroethylene (TCE) on the epididymis and epididymal sperm in mice. Mice were exposed to TCE (1000 ppm) by inhalation for 6 h/day for 5 days/week for 1 to 4 weeks. Segments of the epididymis (caput, corpus and cauda) were examined by light and electron microscopy. At the light microscopic level, degeneration and sloughing of epithelial cells were evident as early as 1 week after TCE exposure, and were most pronounced after 4 weeks. Such epithelial damage was observed in the caput, corpus and cauda regions of the epididymis. Ultrastructural observations revealed vesiculation in the cytoplasm, disintegration of basolateral cell membranes, and sloughing of epithelial cells. Sperm were found in situ in the cytoplasm of degenerated epididymal cells. Additionally, a large number of sperm in the epididymal lumen exhibited abnormalities including malformation of head and tail components. Our results demonstrated that exposure to TCE by inhalation causes damage to the epididymal epithelium and sperm.
Subject(s)
Epididymis/cytology , Epithelial Cells/drug effects , Solvents/toxicity , Spermatozoa/drug effects , Trichloroethylene/toxicity , Administration, Inhalation , Animals , Epithelial Cells/pathology , Epithelial Cells/ultrastructure , Male , Mice , Mice, Inbred Strains , Solvents/administration & dosage , Spermatozoa/pathology , Spermatozoa/ultrastructure , Time Factors , Trichloroethylene/administration & dosageABSTRACT
Lectins were used as probes in conjunction with quantitative analysis to investigate the distribution of different carbohydrate residues in hamster zona pellucida and their possible modification patterns after in vivo fertilization and in vitro egg activation. Several lectins including HPA, WGA, RCA-I, PNA, DSA, BSAIB(4), DBA, AAA and MAA were used to label the zona pellucida of both unfertilized and fertilized eggs. With the exception of PNA and BSAIB(4), the same lectins were also used to label the zona pellucida of oocytes activated in vitro. A multicomparison quantitative analysis of the density of labelling in the inner and outer regions of the zona pellucida before and after fertilization in vivo, as well as after in vitro egg activation, was performed. Of all the lectins studied, preferential localization of labelling by RCA-I and DSA to the inner zona pellucida of unfertilized eggs was observed. After in vivo fertilization, there was an increase in labelling in the inner region of the zona pellucida when thin sections of fertilized oocytes were incubated with HPA, BSAIB(4) and AAA. Although increased labelling by RCA-I was observed, a significant decrease in labelling intensity was obtained with WGA and the sequence Neu-WGA in both the inner and outer zona pellucida of fertilized oocytes. A significant increase in the density of labelling with WGA was also observed after digestion with neuraminidase. In parallel, when hamster oocytes activated in vitro were compared with those fertilized in vivo, a difference in lectin-gold labelling was observed in both the inner and outer region of the zona pellucida. Labelling with HPA, WGA, DSA and MAA increased significantly in both the inner and outer regions of the zona pellucida, whereas labelling by DBA significantly decreased in the inner portion of the zona pellucida. After neuraminidase treatment, a significant increase in labelling density was observed when thin sections of in vitro-activated oocytes were incubated with WGA. These results demonstrate: (i) the post-fertilization modifications of major saccharidic determinants that may play a role in the sperm-egg interaction process of fertilization in vivo; and (ii) that the modified properties of zonae pellucidae of fertilized and in vitro-activated eggs resulting from the action of hydrolytic enzymes, as well as glycoproteins released through exocytosis of cortical granules, are not identical.