Nature of Penicillin-Induced Growth Inhibition of Mycoplasma neurolyticum

When penicillin was added to cultures of Mycoplasma neurolyticum in amounts up to 1,000 units per ml, lag time and generation time were increased and the total population was reduced in proportion to the antibiotic concentration. Although growth suppression by penicillin was complete, the death rate was slow and linear over periods up to 12 days. Growth after induced lag was due to a decay in penicillin activity and was not the result of mutant selection. However, repeated transfer in media which contained increasing concentrations of penicillin resulted in normal growth of M. neurolyticum at penicillin levels as high as 1,000 units per ml. Penicillinase did not play a role in recovery from penicillin-induced lag, and the inactive penicillin molecule did not prevent normal growth of M. neurolyticum. Removal of penicillin from the medium by washing or penicillinase during the induced lag was immediately followed by normal growth of the organism. These results suggest a reversible antibiotic function for penicillin which prevents multiplication of the organism by means unrelated to cell wall formation.     

Lipid composition of Mycoplasma neurolyticum

The total lipid content of Mycoplasma neurolyticum comprises about 14% of the dry weight of the organisms and is about equally distributed between the phospholipid and the neutral-glycolipid fractions. The neutral lipids were identified as triglycerides, diglycerides, and cholesterol. The glycolipid fraction contained 1-O-beta-glucopyranosyl-d-2,3-diglyceride and 1-[O-beta-d-glycopyranosyl-(1-->6)-O-beta-d-glucopyranosyl]-d-2,3-diglyceride. The latter lipid is structurally identical to the diglucosyl diglyceride which occurs in Staphylococcus aureus. The phospholipids of the organism consist of a fully acylated glycerophosphoryl-glycerophosphoryl glycerol, phosphatidic acid, diphosphatidyl glycerol, phosphatidyl glycerol, and amino acyl esters of phosphatidyl glycerol. Phosphatidic acid and phosphatidyl glycerol account for greater than 90% of the phospholipids of organisms in the exponential phase of growth. The predominant fatty acids found in all of the acyl lipids were palmitic, stearic, and oleic acids.     

Growth phases of Mycoplasma in liquid media observed with phase-contrast microscope

Razin, Shmuel (University of Connecticut, Storrs), and Benjamin J. Cosenza. Growth phases of Mycoplasma in liquid media observed with phase-contrast microscope. J. Bacteriol. 91:858-869. 1966-Growth of 11 Mycoplasma strains in liquid media was followed by phase-contrast microscopy. A similar pattern of development was common to all strains. Branching filaments, 0.3 to 0.4 mu thick, characterized the early logarithmic phase of growth. The length of the filaments varied according to the strain tested and the growth medium. Addition of oleic acid to the medium induced the formation of very long filaments by M. laidlawii strain B. Upon aging, the filaments were found to break up into chains of coccoid elements. These chains further fragmented to yield shorter chains and single coccoid elements, which characterized the stationary and decline phases of growth. The size of the coccoid elements increased from 0.3 to 0.4 mu, when formed in the filaments, to 0.6 to 0.8 mu after being released from the chains. Further increase in the size of the cells took place at the decline phase of growth, leading to the formation of very large cells reaching a diameter of 10 to 20 mu. However, these large cells had the appearance of empty vesicles and were apparently nonviable as indicated by viable-count experiments.     

Growth of myxococci in suspension in liquid media

Growth of different strains of myxococci in various liquid media was studied in Erlenmeyer flasks and in small fermentors. Medium containing a small concentration of agar allowed the myxobacteria to grow in the liquid phase in suspension. The dry weight of the cells increased about 100 to 200%. Substitution of other thickening substances for agar caused increased growth with all of eight tested agents.     

Growth of Helicobacter pylori in various liquid and plating media

The objectives of this research were to compare commonly used liquid and plating media to elucidate whether one medium provided superior growth of Helicobacter pylori in vitro. The liquid media compared were Mueller-Hinton broth, brain heart infusion broth and H. pylori special peptone broth, formulated in this laboratory. No significant differences in growth rates were noted and shaking during the incubation of broths was not essential for good growth. The plating media compared included Columbia agar, Mueller-Hinton agar, modified Glupczynski's Brussels campylobacter charcoal agar, Johnson-Murano agar and H. pylori special peptone agar (HPSPA). None of the non-specific plating media that have been used historically to culture H. pylori exhibited any particular advantage. However, HPSPA provided an obvious advantage in colony size. Helicobacter pylori special peptone agar enhances the cultivation of H. pylori and could improve the recovery of the bacterium from clinical samples in vitro.     

Role of liquid versus agar-gelled media in mass propagation and ex vitro survival in bananas

Shoot cultures from meristem explants of three cultivars of banana, ‘Cavendish’, ‘Bluggoe’ and ‘Silk’ were established on Murashige and Skoog medium with 8.9 μM benzyladenine and 0.98 μM indolebutyric acid. Different types of media, such as agar-gelled, agitated liquid and static liquid, were assessed for their ability to support shoot multiplication and ex vitro survival. Liquid media were found better for shoot multiplication whereas agar-gelled medium supported maximum ex vitro survival. For maximum plantlet production, growing in static liquid medium followed by a brief culture on agar-gelled medium has been suggested.     

Mycoplasma Suipneumoniae and Mycoplasma Flocculare in the Growth Precipitation Test

A number of laboratory and field strains of Mycoplasma suipneumoniae and Mycoplasma flocculare were subjected to a comparative examination by the growth precipitation test. It was found that all the strains could readily be identified by that test. Slight evidence of cross-reaction was noted for a few of the laboratory strains, but not until late in the observation period. Only some of the field strains would form precipitates when primary cultures (from tissue suspensions) were used, but all strains could be identified already in the second and third passages. The test therefore seems well suited for distinguishing the two species from each other.     

Growth and sporulation of Entomophthora virulenta on semidefined media in liquid culture

Entomophthora virulenta was tested for growth and sporulation under various nutritional conditions in shake cultures containing combinations of dextrose and protein hydrolysates. Darkness, a temperature of 25°C, and a pH near 6.5 induced the formation of the greatest number of zygospores when the carbon and nitrogen sources were held constant. The influence of the total concentration of nutrients and of the ratio of the carbon and nitrogen sources depended on the nitrogen source used: The highest amount of spores under these conditions was produced with yeast extract as nitrogen source, a carbon/nitrogen source ratio of 2, and a 15% total nutrient concentration. The influence of the preinoculum and inoculum on sporulation is discussed.     

Mycoplasma pneumoniae and Mycoplasma salivarium: Electron Microscopy of Colony Growth in Agar

Colonies of Mycoplasma pneumoniae and Mycoplasma salivarium grown in PPLO agar were examined by light and electron microscopy. The main objective of the investigation was to attempt in situ fixation and minimize tonic changes in the organisms. Microscopy revealed that both organisms grew both in and upon the agar. The agar and surface growths of M. pneumoniae exhibited similar profiles, whereas those of M. salivarium differed strikingly. Both organisms are highly pleomorphic, but their matrix was denser and appeared more intact than in previously reported profiles. Cells which resemble the commonly reported mycoplasma were occasionally observed. The significance of these discrepant profiles remains unanswered. It is suggested that they may represent aged or osmotically damaged cells.     

Growth and comparative physiology ofKlebsiella oxytoca attached to granular activated carbon particles and in liquid media

Experiments were performed to evaluate the comparative growth and physiology ofKlebsiella oxytoca grown attached to granular activated carbon particles (GAC) and in liquid medium. Laboratory studies showed that when this organism attached to GAC, the growth rate was enhanced more than 10 times in the presence of glutamate, a substrate that adsorbed to the surface. No differences were observed if the substrate was glucose, which did not adsorb to GAC. Cellular [³H]thymidine uptake was used to estimate DNA biosynthesis. Attached bacteria grown in a minimal nutrient medium containing 20.0 mg/liter glutamate took up 5 times more [³H]thymidine than cells grown in suspension. [³H]uridine was used as a measure of RNA turnover. Attached cells were shown to assimilate 11 times more [³H]uridine than cells in liquid media. Cell size measurements were performed by differential filtration. Cells grown in a minimal medium with 20.0 mg/liter glutamate decreased in size over time, with 62% of the total number passing through a 1.0m filter after 9 days incubation. In the same period, 39% of a cell population that was grown on GAC passed through a 1.0m filter. These studies indicate that GAC provides an interfacial environment for the enhanced growth ofK. oxytoca when glutamate is the substrate.     

Growth of Mycoplasma mycoides subspecies mycoides on media containing various sugars and amino sugars: an ampoule microcalorimetric study

The growth of Mycoplasma mycoides subspecies mycoides strain T1 on media containing various sugars, tryptose, yeast extract, salts and either pig or calf-serum or a mixture of bovine serum albumin (BSA) plus lipid was followed by ampoule microcalorimetry. Power-time (p-t) curves were reproducible and showed details of growth not observable by conventional microbiological techniques. In media with metabolisable sugars p-t curves typically showed three periods of exponential increase in power separated by transient declines or plateaux. Maximum power (Pmax) was dependent upon the nature and concentration of sugar, whether ampoules were capped in air or nitrogen, and whether the medium contained pig or calf-serum or BSA plus lipid. The highest Pmax was observed in pig-serum medium with glucose, in ampoules capped in air. Decline in power from Pmax was essentially exponential.     

Internalization and intracellular survival of Mycoplasma pneumoniae by non-phagocytic cells

Current theory holds that mycoplasmas remain attached to the surface of epithelial cells although some mycoplasmas have evolved mechanisms for entering host cells that are not naturally phagocytic. The ability of Mycoplasma pneumoniae strain M129 to invade and survive within host cells was studied using a HeLa cell line and a human lung carcinoma cell line (A549). The invasion process into the eukaryotic cells was studied qualitatively by confocal laser scanning microscopy and quantitatively by the gentamicin resistance assay. Internalization was found with A549 cells but not with HeLa cells. Internalization was dependent on the duration of the infection and on temperature. The organism, detected in the cytoplasm and perinuclear regions, survived within the host cells for prolonged periods of time. The intracellular location of M. pneumoniae is obviously a privileged niche, well protected from the immune system and from the action of many antibiotics and may explain the pathogenic potential of this organism.     

Stability of phenylacetic acid in liquid media

The stability of phenylacetic acid (PAA) in H₂O and B5 and MS culture media was determined by HPLC. There was no loss of PAA when a nonsterile 10 mM stock solution was held at 5°C for 2 months. PAA was stable to autoclaving in full-strength MS and B5 media. After storage at 5°C or after agitation at 125 rpm at room temperature for 28 days, 100% of the PAA remained in B5 medium. Under comparable conditions, up to 15% of the PAA was lost in MS medium.     

Growth Factors for Mycoplasma pneumoniae in Yeast and Tissue Extracts

1. Many tissue extracts have been shown to have growth factor activity towards M. pneumoniae notably bovine lung. Yeast extract is therefore not unique in this respect. The materials in lung digest and in yeast extract are very similar in properties and could be identical.
2. The growth factor is a low molecular weight water soluble material. Despite its stability to heat it decomposes on storage, presumably by oxidation.
3. Chromatographic procedures, on both Dowex 50W columns and on paper, indicated the presence of two active components.     

Growth and survival of rumen fungi

The life cycle and growth kinetics of an anaerobic rumen fungus (Neocallimastix R1) in liquid and solid media are described, together with its response to light, temperature and oxygen. These results are discussed in relation to the survival of rumen fungi in saliva and faeces of sheep, and the possible routes for the transfer of anaerobic fungi between ruminants. The thallus and life cycle of Neocallimastix R1 are compared with those of aerobic chytrids.