Ultrastructural alteration of cytolytic T lymphocytes following their interaction with target cells. II. Morphogenesis of secretory granules and intracellular vacuoles.

“Secretory” granules, crystal-like structures, lysosema-like granules, ergastoplasmatic lamellae, electron-opaque ribosomes, and lipid vacuoles were detected in the cytoplasm of cytolytic T lymphocytes. After 30 to 60 min interaction with a target cell the fusion of “secretory” granules and crystal-like structures with lipids, mitochondrion degeneration, hypertrophy and reorientation of the Golgi apparatus toward the contact zone with a target cell were observed.     

Carbohydrate analysis and serological classification of typical and atypical isolates of Aeromonas salmonicida: a rationale for the lipopolysaccharide-based classification of A. salmonicida

The cell envelope of Aeromonas salmonicida contains a lipopolysaccharide (LPS) essential for the physical integrity and functioning of bacterial cell membrane. Using a recently developed in-source fragmentation technique, we screened 39 typical and atypical isolates of A. salmonicida and established their O-chain polysaccharide structure by capillary electrophoresis-mass spectrometry (CE-MS), compositional and linkage analyses and comparison to the previously determined O-chain polysaccharide structure of A. salmonicida strain A449. These studies have demonstrated that A. salmonicida isolates fall into three distinct structural types, types A-C, based on chemical structures of their respective O-chain polysaccharide components. Subsequent immunoblotting and serological studies with salmon polyclonal antisera produced to formalin-fixed cells of A. salmonicida strains A449, N4705 and 33659 representing three structural types A-C revealed that variations in the O-chain polysaccharide structure have led to significant serological differences between strains belonging to type A and non-type A, where non-type A species include chemically separated structural types B and C. Due to the presence of common antigenic determinants shared by their respective O-chain polysaccharide components, serological cross-reactions were observed between A. salmonicida strains belonging to structural types B and C. These findings suggest the possibility of developing LPS-based classification system of A. salmonicida sub-species consisting of two serologically distinct types, type A and non-type A.     

Physical properties of glycosyl diacylglycerols. 2. X-ray diffraction studies of a homologous series of 1,2-Di-O-acyl-3-O-(alpha-D-glucopyranosyl)-sn-glycerols

X-ray diffraction methods were used to characterize the thermotropic polymorphism exhibited by aqueous dispersions of a homologous series of 1,2-O-acyl-3-O-(alpha-D-glucopyranosyl)-sn-glycerols. Upon cooling from temperatures at which the acyl chains of these lipids are melted, all of these compounds form structures that exhibit both low-angle and wide-angle diffraction patterns consistent with the formation of lamellar L beta gel phases. After a suitable protocol of low-temperature annealing, complex diffraction patterns consistent with the formation of highly ordered, lamellar, crystal-like phases are obtained. These patterns are similar for all of the compounds studied, suggesting that the unit cell structure is invariant. The assumption that the unit cell structure is invariant permits the assignment of phases to the diffraction orders, thereby making possible the construction of electron density profiles. These electron density profiles indicate that the crystal-like phases of these lipids are poorly hydrated structures with the hydrocarbon chains inclined at 35 degrees to the bilayer normal. The diffraction patterns of the crystal-like phases of these lipids changed abruptly at the calorimetrically determined phase transition temperatures to those characteristic of either lamellar liquid crystalline phases (N less than or equal to 17) or inverted nonbilayer phases. With these X-ray diffraction data we demonstrate that, at elevated temperatures, the shorter chain homologues (N less than or equal to 16) form cubic phases of the Pn3m space group, whereas the longer chain compounds form inverted hexagonal phases.     

Morphological variation of new Thermoplasma acidophilum isolates from Japanese hot springs.

We isolated 12 strains of Thermoplasma acidophilum from hot springs in Hakone, Japan. T. acidophilum strains showed morphological variation in the crystal-like structure in the cell and the fibrous structure on the cell surface. Two strains tested were sensitive to novobiocin. However, a novobiocin-resistant mutant was obtained by spontaneous mutation.     

Antigenic structure of saprophytic leptospirae and their classification]

The authors present the results of study of the serological properties of 46 strains of saprophytic leptospirae of different origin. On the basis of the affinity of the antigenic structure detected in the cross microagglutination reaction (MAR) 38 strains under study were united into 8 serological groups; the rest 8 strains were serologically independent in this reaction. The fact that 9 strains of water leptospirae isolated in the Armenian SSR belonged to one serological group was proved in the cross MAR and the test of aglutinin adsorption. This serological group was new and was named L. armenica. Five individual serological types of saprophytic leptospirae were differentiated in its composition. Comparative study of the serological interrelations between the group of strains isolated in Armenia and the strains of some serological groups and serological types the closest serological connections were noted in the K-1030 (serological group Armenica) and Bovedo (serological group Andamana) strains. It is believed that the existing division of the saprophytic leptospirae into two serological groups (Semaranga and Andamana) required widening and supplement by new serological groups and serological types.     

Distribution and Properties of Serological Types of Streptococcus faecium, Streptococcus durans and Related Strains

S ummary . The distribution of 19 serological types of Streptococcus faecium and related organisms has been studied, using 367 strains isolated mainly from faecal samples. Several types occurred in man as well as in pigs, sheep, cattle or chickens.
Strains of the same serological type showed a diversity of fermentation reactions, so that organisms which could be identified as Streptococcus durans shared a common type antigen with Strep. faecium strains. The evidence given here supports the proposal that Strep. durans should in future be considered as a variant of Strep. faecium .
It has also been shown that, in common with those of Streptococcus faecalis , the type antigens of the Strep. faecium types are cell wall components.     

Ultrastructure of the conjugates of cytolytic T-lymphocytes and target cells]

Conjugates of target cells and of cytological T-lymphocytes obtained on the 11th day after alloimmunization were investigated. The conjugates formed small and medium lymphocytes; mature secretory granules, crystal-like structures and lipids were revealed in their cytoplasm. The lymphocyte is spherical, the area of contact with the target cell does not exceed 5 to 15%. Cytolysis of target cells is observed after 30 to 60 minutes of incubation. The lymphocyte becomes flattened, its nucleus acquires an oval form, and the area of contact with the target cell increases considerably. At the same time hypertrophy and change of orientation of Golgi's complex to the area of contact with the target cell, coalescence of the secretory granules with lipids and crystal-like structures, the appearance of immature secretory granules and vacuolar degeneration of mitochondria are demonstrated. The lymphocyte membrane becomes "desquamated"; structures connected with it, named "membranosomes" are described. It is suggested that the secretory processes in the cytoplasm of cytolytic T-lymphocytes are activated in their interaction with target cells.     

Comparative study of the antibiotic sensitivity of Proteus hauseri bacteria belonging to different serological groups]

Sensitivity to 9 antibiotics of 1040 strains of Proteus belonging to the serological groups 03, 05, 06, 07, 010, 011, 013, 023, 024, 026, 027, 028 and 030 was studied. It was found that the above strains were sensitive and highly sensitive to the aminoglycosides and streptomycin, slightly sensitive to levomycetin and resistant to tetracyclines, erythromycin and penicillin. All the strains were polyresistant and 99.6 per cent of them were resistant to 4--9 antibiotics. Ten types of resistance were found. Proteus strains with the resistance type PETOtCht were most common. No relation between the occurrence of the strains of various serological groups and the character and level of their resistance to the antibiotics was found.     

Molecular self-organization and multiple equilibrium systems

We have compared different types of natural self-organizing systems: crystal-like formation of multimolecular systems were conferred with self-organization in active media and dissipative structure formation. The comparison revealed a common feature of all such systems. They all have bi-(multi-)stable states. We propose a hypothesis that self-organization is impossible in systems that could not have bi-(multi-)stable states.     

Deoxyribonucleic Acid Base Composition and Homology Studies of Leptospira

Four distinct genetic groups of leptospiras were demonstrated among selected pathogenic and "biflexa" serological types. Pathogenic leptospiras could be divided into two groups on the basis of per cent guanine + cytosine (GC) in their deoxyribonucleic acid (DNA). One group had 36 +/- 1%, the other 39 +/- 1%. The biflexa strains had DNA of 39 +/- 1% GC, but were further separated into two groups on the basis of DNA-annealing tests. Strains within groups had a high degree of specific duplex formation (75% binding or more with reference to the homologous DNA). There was little or no genetic relatedness between strains of the four groups (less than 10% DNA homology). The thermal elution midpoint of heterologous DNA duplexes was always lower than the homologous reaction. The serological relationships among strains were not meaningful in terms of relatedness determined by specific duplex formation.     

Characterization of thermosensitive mutants of Physarum polycephalum

Summary Amoebal thermosensitive mutants of Physarum polycephalum have been isolated after mutagenesis of the amoebal form by nitrosoguanidine treatment. About 70% of the independent thermosensitive amoebal mutants obtained were also thermosensitive in the plasmodial form. Two basic screening methods were applied at the same time to thermosensitive microplasmodia in order to detect strains defective in premitotic events, mitosis or chromosomal DNA synthesis. The first method consists in the determination of increase in protein. RNA and DNA with incubation time at the non-permissive temperature. It allowed the detection of four independent thermosensitive mutant strains, showing an early arrest in DNA synthesis. The second one is the quantification of the variations of the different nuclear types at the restrictive temperature. Two mutant strains presented very large nuclei, uni- or multinucleolate, very similar to those obtained after methyl benzimidazole carbamate treatment, suggesting a defect in one of the mitotic processes. One of these two mutant strains showed an early arrest in DNA synthesis at the restrictive temperature. These two screening procedures were completed by electron microscopic observation. This technique allowed the detection of intra-nuclear macrotubular crystal-like structures in a thermosensitive mutant showing a reduced DNA synthesis at the non-permissive temperature.     

A Light and Transmission Electron Microscope Study of a Black Locust Tree, Robinia pseudoacacia (Fabaceae), Affected by Witches'-Broom, and Classification of the Associated Phytoplasma

Light and transmission electron microscopy of phloem sieve-tube elements, companion cells, and parenchymal cells in thin and ultrathin sections of small and medium rachises and small, medium and large leaflets of a black locust tree, Robinia pseudoacacia L., affected by witches'-broom disease revealed (in the small and medium rachises and leaflets) structures that were characteristic of phytoplasmas, and crystal-like inclusions in the phloem sieve-tube members. A crystal-like inclusion was also seen in a companion cell. Paracrystalline arrays were seen only (and very rarely) in phloem sieve-tube elements of medium rachises. Some elements contained several crystal-like inclusions and each inclusion had fracture planes. The crystal-like inclusions and paracrystalline arrays apparently have not been previously reported in the black locust. The paracrystalline arrays and crystal-like inclusions may merely be by-products of the plant's metabolic activity. Extensive additional work would be required to establish the precise relationship (if any) of the arrays and inclusions to the black locust, witches'-brooming and/or phytoplasmas. Results from analysis of 16S rRNA gene sequences amplified by the polymerase chain reaction indicated for the first time that the phytoplasma associated with black locust witches'-broom is a member of group 16 SrIII (peach X-disease) phytoplasma group. This raised the question of whether black locust may be a significant source of the phytoplasma for infection of other plants, especially agricultural crops.     

Determination of cell wall teichoic acid structure of staphylococci by rapid chemical and serological screening methods

Investigations of cell wall teichoic acid structures of various staphylococci were carried out by a rapid method based on the gas-liquid chromatographic separation of products obtained after treatment of phenol-extracted cells with 70% hydrofluoric acid. In most of the strains teichoic acids of the poly(glycerolphosphate) and/or poly(ribitolphosphate) type were found. Teichoic acids of the poly(glycerolphosphate-N-acetylglucosaminephosphate) type and polymers consisting of N-acetylglucosaminephosphate were present in few strains.The results obtained by the rapid chemical screening method were compared with data obtained by serological analysis of teichoic acid structures using specific antisera and the lectin wheat germ agglutinin. Teichoic acid components occurring in low concentrations could only be detected with the chemical and not with the serological method. A number of strains of species of the genus Staphylococcus have been studied using these rapid methods. With a few exceptions, the teichoic acid structure proved to be a constant marker within a given species.Abbreviations used CIE counnter immunoelectrophoresis - GalNAc N-acetylgalactosamine - Glc glucose - GlcNAc N-acetylglucosamine - Gro glycerol - Rit ribitol     

SEROLOGICAL STUDIES OF MALE AND FEMALE STRAINS OF EUDORINA ELEGANS (CHLOROPHYCEAE)1,2

Male and female strains of Eudorina elegans Ehrenberg were examined by serological methods for flagellar and cellular antigenic differences. No significant differences could be detected between the strains by using agglutination, immobilization and diffusion tests. However, differentiated sperm were neither agglutinated nor immobilized by anti-vegetative cell sera.     

Biochemical typing as a method of differentiating group B streptococcus]

In epidemiological studies on the group B streptococcus the serological typing is used. The paper present the results of a study on usefulness of biochemical typing for differentiation of the group B streptococcus. For that purpose, 210 strains descended from colonized infants and pregnant women were put to typing with both of mentioned methods. We showed that each of the method distinguishes similar number of biotypes and serotypes. However, ought to be marked that significant number of strains (93.8%) belonged to the three out of eight biochemical types. Similar results were achieved in serological typing, three of the most numerous serotypes contained 81.4% strains. Analysis of the relationship between serological and biochemical types did not reveal statistical association because the strains belonged to various serotypes. Obtained results show that both methods of typing--biochemical and serological, have similar value in differentiation of the strains. The method of biochemical typing is quite simple and can be used in laboratory conditions.     

Experience with epidemiologic studies of pyocyaneus infections. I. Feasibility of using serotypes and antibiotic resistance as a epidemiologic label for clinical strains]

The authors carried out serological typing of 98 Pseudomonas aeruginosa strains, isolated from patients of burn department of the Sklifosovsky First Aid Institute in January-July, 1974, and of 215 strains obtained from other sources; their sensitivity to 13 antibiotics was determined. Pseudomonas aeruginosa cultures isolated from the patients were typed with O-sera of 10 serological types. The presence of several hospital strains of Pseudomonas aeruginosa was found by means of serological typing; along with these there were revealed cultures of this causative agent sporadically appearing in the department. Sensitivity to some antibiotics could serve as an additional criterion for differentiation of Pseudomonas aeruginosa strains of the same serological type.     

Serologic properties of strains of Cl. perfringens isolated from the intestinal contents of healthy subjects]

A study was made of the quantitative content in the intestine of C1. perfringens strains in 6 healthy persons who stayed in a hermetically sealed space for 1 month and for 1 year. C1. perfingens strains were isolated from the fecal samples of each of the volunteers at various periods of the trial. A total of 570 strains of C1. perfringens of type A with anticellular sera obtained to the strains of various serological groups were studied. Serological properties of C1. perfringens strains of type A present in the intestinal contents of man were nonhomogeneous. This pointed to the simultaneous presence in the intestine of strains belonging to several serological types. A partial or complete replacement of one strain by another (differing by serological properties) occurred in the course of not over one month. C1. perfringens strains of type A present in the intestine of each volunteer were subdivided into serological types individual for each of the persons under observation. This pointed to the fact than no interexchange of strains of the mentioned bacteria occurred between different persons in the hermetically sealed space.     

Antigenic properties of genetic recombinants of serologically typed E. coli]

The conjugation between the typed strains of E. coli belonging to various serological groups and conjugation between typed and untyped E. coli strains were studied. Genetic determinant controlling the synthesis of the O100 antigen proved to be closely linked with histidine locus. Among recombinants obtained in crossing the typed E. coli strains there were such belonging to the serological type different from the serological types of donor and recipient cells.     

Role of horizontal genetic exchange in the antigenic variation of the class 1 outer membrane protein of Neisseria meningitidis

The nucleotide sequences of the genes encoding the class 1 outer membrane protein of Neisseria meningitidis (PorA) from 15 meningococcal isolates have been examined. These strains, isolated over a number of years, represented a variety of serological types, clonal groups, and geographical locations. Analysis of the aligned nucleotide sequences showed that the known serological relationships between these proteins were not necessarily reflected throughout the nucleotide sequences of their genes. The uneven distribution of base substitutions, revealed by a comparison of the informative bases, suggested that these genes possessed a mosaic structure. This structure probably resulted from the horizontal transfer of DNA between strains and would have contributed to both the generation and the spread of novel antigenic variants of the protein. In addition, the nucleotide differences between porA genes from different strains were not consistent with the nucleotide sequence divergence of the whole chromosome, as indicated by pulsed-field gel electrophoresis (PFGE) fingerprinting techniques: some strains with divergent PFGE fingerprints shared porA genes with extensive regions of nucleotide sequence identity and, conversely, some strains with similar chromosome structures possessed porA genes with different nucleotide sequences and serological properties. This suggested that entire genes had been exchanged between strains. Given that the meningococcal class 1 OMP is a major component in novel vaccines, some of which are currently undergoing field trials, the potential of horizontal genetic exchange to generate antigenic diversity has implications for the design of such vaccines.     

Investigations on the antigenic structure of actinomycetales IX. Serological classification of the Nocardiae with the polysaccharide fractions of their cell walls

Summary Polysaccharide fractions were extracted with 7.5% sulphosalicylic acid from defatted cell walls of 43 strains of nocardiae. All the polysaccharides, except those ofN. turbata, revealed chromatographically glucosamine, galactose, glucose and arabinose. Rhamnose was only found in the fractions isolated from some strains.The complement-fixation test with the polysaccharide fractions and anti-cell-wall sera allowed to distinguish four serological groups and 21 types among the strains examined. A method and a pattern of the serological classification of nocardiae has been presented.This work was partly conducted under a Von Humboldt Associate-Professor Fellowship in the Hygiene-Institute, University of Bonn, in 1963.Part of this paper was read at the meeting of the International Society of Human and Animal Mycology during the International Botanical Congress in Edinburgh, Scotland, 1964.