CALIFORNIA STATE UNIVERSITY, NORTHRIDGE PATTERNS OF GROWTH AND GLIDING ~10TILITY !t OF SIMONSIELLA AND ALYSIELLA A thesis submitted in partial satisfaction of the requirements for the degree of Master of Science in Biology by Gordon Emery ____Buchanan, ,/ Jr. January, 1976 The thesis of Gordon Emery Buchanan, Jr. is approved: California State University, Northridge December, 1975 ii ACKNOWLEDGt4ENTS I am especially indebted to the members of my graduate committee, :or. Daisy A. Kuhn, Dr. Marvin H. Cantor and Dr. Charles R. Spotts, who :helped me in manY ways during my years as a graduate student at California State University, Northridge. I am thankful for the continual assistance and encouragement from my colleagues, David Gregory, Roland Gunther and Christie Jenkins, who also participated in the research on the taxonomy of Simonsiella and ~siella. I also acknowledge gratefully the help and advice given to me by Carla Bowman, Ruth Jung, David Oakland, Flora Price, Bernardine Pregerson, and Edward Rau. iii TABLE OF CONTENTS Page List of Tables.................................................... v List of Figures. vi Abstract. .vii Introduction ......... • • • • • • • • • • • • • • • • • • • • • • • • • • • • ot ••••••••••••••• 1 Methods and Materials. 7 Results and Discussion. 17 Conclusions ........... .. 76 References . ........ - . 80 iv LIST OF TABLES Page Table 1. Colony characteristics of Simonsiellaceae ................. l8 Table 2. Types of patterns of growth and gliding motility.; ........ 29 Table 3. Rates of gliding motility ..................•.............. 36 Table 4. Effect of temperature on growth ........................... 41 Table 5. Effect of temperature on patterns of growth and gliding motility .........................•................ 43 .Table6. Effect of pH on growth .....................•.... ~ ......... 46 Table 7. Effect of pH on patterns of growth and gliding motility ................•..........................•...... 47 Table 8. Effect of NaCl on growth .................................. 48 Table 9. Effect of NaCl on patterns of growth and gliding mot i 1i ty . .. ·. .. 51 Table 10. Effect of 11 Flexibacteria Medium 111 on growth .......•...... 53 , Table 11. Effect of 11 Flexibacteria Medium 111 on patterns of growth and gl i d1 ng moti1 i ty ............................... 55 Table 12. Effect of further modifications of 11 Flexibacteria Medi urn 1 11 on growth ....................................... 56 :Table 13. Effect of further modifications of 11 Flexibacteria Medium 111 on patterns of growth and gliding motility ...... 58 Table 14. Effect of omission of salts from BSTSY on growth .......... 60 'Table 15. Effect of omission of salts from BSTSY on patterns of growth and gliding moti 1i ty ............................ 61 Table 16. Effect of humidity on grm.,rth and gliding motility. I. .... 63 Table 17. Effect of humidity on growth and gliding motility. !! .... 64 Table 18. Effect of agar concentration on growth .................... 65 Table 19. Effect of agar concentration on patterns of growth and gliding motility ...................................... 66 Table 20. Effect of used BSTSY broth on growth. 1 ....•............. 69 v Page Table 21. Effect of used BSTSY broth on patterns of growth and gliding motility. I. ................................. 70 Table 22. Effect of used BSTSY broth on growth. !! ................. 72 Table 23. Effect of used BSTSY broth on patterns of growth and g1 i d i ng mot i1 i ty . I I. 73 LIST OF FIGURES Figure 1. Colony morphology types of dog Simonsiella strains ........ 22 Figure 2. Colony morphology types of human Simonsiella strains ................................................... 24 ,Figure 3. Colony morphology types of sheep Simonsiella strains ........................•.......................... 26 Figure 4. Colony morphology types of cat Simonsiella strains ........ 28 vi ABSTRACT PATTERNS OF GROWTH AND GLIDING MOTILITY OF SIMONSIELLA AND ALYSIELLA by Gordon Emery Buchanan, Jr. Master of Science in Biology January, 1976 Forty six strains of Simonsiella and two strains of Alysiella, large filamentous, gliding bacteria from the oral cavities of warm- blooded animals, were grown under various environmental conditions in .order to study aspects of the taxonomy of these organisms. Differences in growth with changes in temperature, pH, and NaCl and synthetic sea water salts concentrations segregated the strains into several :individual groups which correlated with the isolation of the strains from dogs, sheep or humans. These findings add to the information derived from studies on the cellular morphology and on the biochemistry :and physiology (Kuhn et ~·, 1974; Pangborn et al_., 1973 and 1974; . Nyby, 1974; Gregory, 1975), on the fatty acid profiles (Jenkins, 1976), and on the mole percent G+C content (Kuhn et .!!_., 1974). Microscopic features of gliding motility and growth patterns of Simonsiellaceae are recorded for the first time. Growth on agar viewed microscopically revealed that a small fraction of Simonsiellaceae . strains glided, whereas other strains displayed types of colony vii morphology that ranged from entire-edged to pronounced filamentous growth. Gliding motility rates were found to range from 5 llm/min ; to 23.8 llm/min. Gliding motility was manifested by individual, frequently well-separated fi 1aments rather than by 11 armi es 11 of closely-associated cells as described by Stanier (1942a) for the cytophagas. Gliding motility appeared to occur as _necessary to ·reach areas on the agar surface supplying fresh nutrients, and ,when these areas were reached gliding motility was arrested and :secondary colony formation ensued. Gliding motility was frequently most pronounced in regions of heavy growth bordering unoccupied agar ·surfaces, suggesting that filaments glide in response to gradients of either decreasing waste metabolites or increasing fresh nutrients. The influence of other factors such as light, agar concentration and 'humid incubation are considered. viii ~~- --- ·-· -·· - - -·· ·· · · -~ NTROilucnoN ·· ···· ····- -- ·-· · - ~~ -- - - - ···· ' 1.' ! 1 i / Observations of gliding motility among procaryotic organisms were 1reported as early as the 18th century by botanists studying the morph- 1 !ology and taxonomy of the cyanobacteria (Burkholder, 1934). Experi­ i ! :mental investigations of gliding motility began in the late 19th : :century. Unusual aspects of gliding motility such as the swarming of i i"armies 11 of cells (Stanier, 1942a) in the cytophagas and in the ! :myxobacteria, and mucilaginous sheath production and axial rotation of ; i ~gliding filaments in Oscillatoria (Burkholder, 1934; Halfen and :castenholz, 1970 and 1971) have been described. Theories for the 'mechanism of gliding motility have been proposed based on (1) osmotic or surface tension gradients along gliding filaments (Burkholder, ;1934); (2) slime production (Burkholder, 1934; Dodd, 1960), and (3) :rhythmic contractile waves (Jarosch, 1962; Doetsch and Hageage, 1968). -- i ' ';Energy requirements for gliding motility have been calculated (Ha1fen .and Castenholz, 1971). Researchers have observed with transmission electron microscopy and freeze-fracture techniques cell wall features possibly related to the structural apparatus responsible for gliding :motility (Pate and Ordal, 1967; Halfen and Castenholz, 1970 and 19-71; 1Burchard and Brown, 1973). However, in spite of experimental work on 'gliding motility, the basic cellular mechanisms involved remain 'largely a mystery (Doetsch and Hageage, 1968). Vets certain observed !features of gliding motility can be summarized. Gliding motility is found among some species of both the bacteria land the cyanobacteria, the two groups of the Kingdom Procaryotae, as I joutlined in Bergey•s Manual of Determinative Bacter-iology, Eighth L_~ 1 2 :Edition (Buchanan and Gibbons, 1974). These procaryotic gliding 'organisms may be divided into the following groups: (1) the unicellular and multicellular, filamentous cyanobacteria, represented by the orders Chroococcales and Hormogonales, respectively (Desikachary, 1973); (2) the fruiting myxobacteria of the order Myxobacterales; and (3) all other non-fruiting unicellular and multicellular, filamentous bacteria composing the order Cytophagales which includes the families Cyto­ phagaceae, Beggiatoaceae, Simonsiellaceae, Leucotrichaceae and, possibly, Pelonemataceae and Achromatiaceae (Soriano and Lewin, 1965; Buchanan and Gibbons, 1974). Gliding motility may occur among some or all members of each group, or, occasionally, may be limited to certain juvenile or reproductive forms such as the hormogonia of some cyanobacteria, or the conidia or gonidia of Thiothrix and Leucothrix, respectively (Doetsch and Hageage, 1968). Gliding motility is defined as an active movement of an organism in contact with a solid substratum where there is neither a visible organ responsible for the movement nor a distinct change in the shape of the organism (Jarosch, 1962). Gliding motility among some filamentous cyanobacteria may occur even when a filament makes contact with a substrate along only a fraction of its length (Stanier, l942b), or may occur within medium solidified with 0.5% to greater than 3.5% agar (Castenholz, 1973). Gliding organisms placed in a liquid medium have occasionally shown darting, jerking or rolling motions, particularly after making contact with the side of the vessel (Doetsch and Hageage, 1968). Other workers have observed a bending or flexing of the apical portions