- Volume 44, Issue 1, 1966

Summary: This paper presents data to show that loss of viability of Escherichia coli and phage in an aerosol was the consequence of at least three mechanisms. Phage T7 was used and shown to be unstable when sprayed into air. When previously adsorbed to E. coli the aerosol stability of phage T7 appeared to be dependent upon its stage of development in its host. The host E. coli strain B in an aerosol was itself subject to two stresses which operated through different mechanisms within the bacterium: (i) a relative humidity stress having its locus of action such that colony formation and the production of phage T7 were both impaired; (ii) an air stress such that colony formation was impaired but that the bacterium was able to produce phage T7.

Summary: The morphology of seven temperate phages from lysogenic strains of Alcaligenes faecalis and two virulent phages obtained from sewage and active on this species was examined by a negative-staining technique. The phages showed a variety of morphological forms and two of them have unusual combinations of features. One (A11/A79) has a contractile tail sheath around a hollow core, but lacks the base plate, pins and tail fibres usually associated with this type of tail. The other (A5/A6) resembles the C1, F1 group of coliphages but has an octahedral head and a collar. The five remaining temperate phages are similar to the C1, F1 and Providence 9000/9402 group of phages. The sewage phages differ in many respects from one another and also from the temperate phages examined. At least three of the nine phages have octahedral heads.

Summary: Seven flagella-shape mutants were isolated from a curly-flagella strain of Salmonella abortusequi. The motility in broth and spreading ability on semi-solid medium of these mutants as well as flagellar morphology were examined by dark field and electron microscopy. They were classified into the following five mutant types, heteromorphous, small-amplitude, para-curly, short and hooked-curly.

Each of these mutants manifests its characteristic flagellar shape and motility, and the spreading ability of these mutants on semi-solid medium decreases in the order: normal, heteromorphous, small-amplitude, para-curly, short, hooked-curly, curly.

Except in the small-amplitude and short mutants, the shape of a flagellar bundle of living bacteria under the dark-field microscope corresponded in each mutant to a spiral of the flagella observed under the electron microscope. In the small-amplitude mutant, transconformation of the flagellar bundle from small-amplitude to curly was observed in organisms suspended in 0.5% (w/v) methylcellulose solution. In the short mutant, the flagellar bundle was not seen in the dark-field microscope.

Difference of antigenic specificities between the flagella of the parental curly and each of the mutants were not detected when examined by absorption-agglutination tests.

From transduction analyses with P 22 phage, it was found that the traits of each of the four mutants heteromorphous, small-amplitude, para-curly, short were transferred with the structural gene of phase-2 flagellin and had no effect on the flagellar shape and motility in phase-1. From this, it is inferred that the characteristic flagellar shape and motility of these mutants are primarily to be attributed to the conformation of the flagellin which composes the mutant flagella.

Summary: A new species of the genus Bacillus was isolated from Macquarie Island soil. The organism is psychrotrophic, producing spores at temperatures down to and including 0°. The maximum temperature for growth is 25°. Apart from its temperature relations, it most closely resembles B. pulvifaciens, but differs from that organism in its ability to produce acid from carbohydrates with ammonium salts as sole nitrogen source and in its inability to hydrolyse casein or gelatin, to grow on soybean agar or to grow in 5% NaCl. The name Bacillus macquariensis is proposed.

Vegetative organisms stained Gram-negative at all stages of growth. Electron micrographs revealed cell-wall structures typical of Gram-positive bacteria; on the other hand, chemical analyses demonstrated in cell-wall preparations a wide range of amino acids and relatively low amounts of amino sugars, as commonly found in Gram-negative species.

Summary: The BHK21-C13 line of hamster fibroblasts was infected with human parainfluenza virus type 2. The infection persisted through 40 subcultures without affecting the plating efficiency of the cells and without degeneration of the monolayers. Although all cells contained fluorescent-staining virus antigen and most became specifically haemadsorbing and contained inclusion bodies, the titre of virus in 4-day-old cultures was only equivalent to 0.0001 plaque-forming unit/cell. Infection of cells continued when clones were grown in anti-parainfluenzal serum. There was only very slight interference to infection with other viruses and superinfection with parainfluenza type 2 virus caused degenration of the culture. Persistent infection of BHK21 cells with human type 2 parainfluenza virus producedno loss of contact inhibition and no ability to grow in semi-solid media, changes which are characteristic of BHK21 cells when transformed by polyoma virus of Rous sarcoma virus.

Summary: Immunoelectrophoretic analysis (IEA) of cytoplasmic proteins of Neurospora crassa revealed more than 30 antigenic components in detectable concentrations in crude extracts. The IEA patterns varied somewhat with culture conditions and age, and with strain. A one-gene polymorphism was recognized but was not associated with any known bio-chemical or morphological mutant character. Comparison of the IEA patterns given by proteins of several species of fungi with antisera against N. crassa revealed the prominent phyletic affinities among the protein antigens within the class Ascomycetes. The results are discussed with emphasis on the methods and the interpretation of immunoelectrophoretic variability.

Summary: An Actinoplanes sp. (Actinomycetales) isolated from Blelham Tarn is described and illustrated. It is characterized by a filed arrangement of spores both in sporangia and conidiophore systems, suggesting that these two types of reproductive structure may develop in an essentially similar manner.

Summary: Aconitase and NAD- and NADP-linked isocitric dehydrogenases were examined in two strains of Aspergillus niger. The mutant strain 72-44 produced higher yields of citric acid on the culture medium used than did the parent strain 72-4. After growth for 22 hr on a medium in which citric acid did not accumulate, the amounts of each enzyme in the parent strain were approximately the same as those in the mutant strain. The enzymes were also found, though in lower amounts, throughout the incubation period of 8 days in both strains grown on a medium in which citric acid accumulated. A 20-fold increase in the concentration of iron in this medium doubled the activity of aconitase in extracts from the mutant strain, though citric acid accumulation was only decreased by 25%. The addition of monofluoro-acetate to the culture medium was toxic to the mould and did not stimulate citric acid yields. It is suggested that during the incubation, some recycling of citric acid may take place. The difference in citric acid yields from the parent and mutant strains was not accounted for on the basis of the degrees of aconitase or isocitric dehydrogenase activity.

Summary: The infrared absorption spectra of forty strains of sulphate-reducing bacteria have been recorded over the range 1800--800 cm.-1. It was possible to identify positively Desulfovibrio gigas and the thermophilic Desulfotomaculum nigrificans. A broad differentiation was possible between salt-water and fresh-water forms of Desulfovibrio desulfuricans. Desulfotomaculum orientis and D. ruminis showed few characteristic differences from the fresh-water strains of D. desulfuricans but there was an indication that, with a more refined technique, at least one of the features otherwise characteristic of the other (thermophilic) desulfotomaculum species might be discerned. The results support to a considerable extent current views on the classification ofthe sulphate-reducers.

Summary: An investigation of the effects on Paramecium aurelia of crude and a highly purified tetanus neurotoxin preparation free from oxidase and haemolysin activity was made to assess the value of this organism in a bioassay system and as a convenient organism for the study of the mode of action of the toxin. Whereas crude tetanus toxin produced cytotoxic changes in P. aurelia, purified tetanus neurotoxin did not even when the paramecia were exposed to ten human lethal doses.

Incubation of the paramecia with fluorescent-labelled toxin revealed that it accumulated within food vacuoles and in focal areas of cytoplasm but not in the cell membrane. Cytologic studies of the paramecia by means of light and electron microscopy revealed no abnormalities.

Summary: Attempts have been made to use the protein composition of streptomycetes as a criterion in their classification. By using polyacrylamide gel electrophoresis, negatively charged proteins in extracts of various streptomycete strains were separated. For each strain a distinctive pattern of protein bands was obtained. Protein bands were mapped for a number of strains, including several each of Streptomyces venezuelae and S. griseus. Maps were compared and contrasted: in general there was more similarity between strains within one species than between strains of different species. Although the polyacrylamide gel technique cannot yet be used as a major criterion in classification, it can serve, in conjunction with other criteria, as an aid in streptomycete taxonomy.

Summary: The cell walls of six different species of algae were prepared in highly purified form and their amino acid and amino sugar contents determined qualitatively. A limited number of ninhydrin-positive components was found, implying that the walls contain peptide rather than protein. In five of the algal walls, those of the Chlorella species, a Scenedesmus and a Lyngbya, there were eight amino acids found in common: aspartic acid, glutamic acid, glycine, alanine, serine, valine, leucine, isoleucine. In addition to these common amino acids, the walls contained sometimes proline or hydroxyproline and different combinations of amino sugar which were characteristic of the species. In the sixth algal wall preparation, that of Nostoc, only five of the eight common amino acids were found along with hydroxyproline. Muramic acid and diaminopimelic acid were found in Lyngbya walls but in none of the others, excepting Nostoc walls which may have contained small amounts of diaminopimelic acid. Carotenoids were found associated with crude wall preparations of som of these algae.The above is taken as evidence for the existence of wall peptide in algae

Summary: The effect of polyene antibiotics upon growth and sterol-induction of oospore production by Pythium periplocum was studied. Filipin and fungichromin markedly inhibited growth and prevented reproduction. Amphotericin B also prevented oospore production but only slightly inhibited growth. Pimaricin slightly inhibited growth and greatly decreased reproduction. Nystatin usually had no effect on growth and slightly stimulated reproduction. At 4 μg./ml., amphotericin B and fungichromin prevented oospore induction by 1 μg. cholesterol/ml. but had no effect on growth. Growth inhibition by amphotericin B and fungichromin was almost completely annulled by higher cholesterol concentrations, and prevention of oospore formation was partially annulled. These results generally support the hypothesis that polyene antibiotics act by interfering with the action of cellular sterols, but the growth inhibition which occurred on sterol-free medium suggests that a second mechanism may be operative.

Summary: Nine spontaneous mutants with altered forms of flagellar antigen i were obtained by picking more rapidly spreading swarms from growth in semi-solid medium containing enough anti-i serum to retard spreading growth. One mutant was in a line of Salmonella typhi given antigen i by transduction, the rest in S. typhimurium strain LT2 adeC-7 proA-46. Two mutants of independent origin were serologically identical and presumably arose by a recurrence of the same mutation. Bacteria expressing the mutant phase-1 antigen were normally motile and the LT2 mutants showed normal phase-variation, to give cultures with an apparently unaltered phase-2 antigen, 1,2,3. Flagellate bacteria with flagella of two of the mutant types, iM6 and iM9, were agglutinated to titres 8--16 by sera from uninoculated rabbits and to titres 50--100 by sera from rabbits immunized with unrelated antigens; suspensions of flagella of these types, but not of others, caused flocculation of indian ink.

The residual activity of anti-i (wild type) sera fully absorbed with mutant antigens showed that each mutant antigen had lost some of the serological specificity of the wild-type antigen; the complex pattern of residual activity on mutant and wild-type antigens of anti-i (wild type) sera absorbed with pairs of mutant antigens indicated the existence of at least 13 antigenic factors in the wild-type antigen, and that each ofthe serologically distinct mutant antigens lacked a different combination of these factors.

The residual activities on the homologous antigens of antimutant sera fully absorbed with wild-type antigen showed that all the mutant antigens, except perhaps iM6 and iM9, had antigenic specificities absent from the wild-type antigen. Each of the 8 serologically different antigens had a unique new specificity, but antigens iM7, iM10 and iM12 shared some new factors.

Attempts to infer the linear order of the presumed sites of amino acid substitution in the polypeptide chain of flagellin from the serological data were unsuccessful; this probably indicates the incorrectness of an assumption involved: namely, that anti-flagellar antibodies have an absolute affinity for, and only for, all of the amino acid side-chains (or all of a reactive subset of them) in a relevant length of polypeptide chain.

Summary: L forms were produced by the penicillin gradient plate technique from four strains of Neisseria meningitidis. All strains were group B and had various sulphadiazine sensitivities. One parent strain had been propagated for many years in the laboratory, whereas the other three were isolated from recent cases of meningitis. To date, each L form has had more than 60 serial passages on medium containing penicillin. From two strains stabilized L form variants developed on penicillin-free medium following 30 or 40 such passages. Morphological characteristics of these organisms were similar to L forms of other bacteria. Medium and environmental conditions necessary for optimal growth included: brain heart infusion containing a final agar concentration of 1.2% (w/v), sucrose, 10% (w/v), horse serum, 10% (v/v), pH 7.4, temperature 37° and increased CO2 tension (candle jar). The L forms were non-groupable and had various fermentative reactions, whereas their sensitivity to sulphadiazine was the same as that of their respective parents. Revertants were produced on penicillin-free medium following every tenth serial passage in the L-form state. Sensitivities to sulphadiazine, fermentative reactions and serological groups of the revertant strains were identical with those of their respective parents.