A Preliminary Study of the Algicidal Mechanism of Bioactive Metabolites ofBrevibacillus laterosporusonOscillatoriain Prawn Ponds

The algae,Oscillatoria, is commonly found in prawn ponds and can lead to reduced productivity. We examined metabolites of the bacteriaBrevibacillus laterosporusfor algicidal qualities. To determine the possible algicidal mechanisms of these bioactive metabolites, different amounts of sterile filtrate of bacterial suspensions were added to cultures containingOscillatoria. The dry weight, the concentrations of chlorophyll-a (chl-a), phycobiliprotein (PC, phycocyanin; APC, allophycocyanin; PE, phycoerythrin), and MDA (malondialdehyde) and the activities of SOD (superoxide dismutase), POD (peroxidase), and CAT (catalase) of algae were measured during the algicidal application. The results showed that lower concentrations of the sterile filtrate (addition ≤ 4 mL) accelerated the growth rate ofOscillatoria, but significant inhibition and lysis were observed with higher concentrations (addition ≥ 8 mL). In two trials (the additions were 8 mL and 10 mL, respectively), the algal dry weights were reduced by 26.02% and 45.30%, and the chl-a concentrations were decreased by 46.88% and 63.73%, respectively, after seven days. During the algicidal treatment, the concentrations of PC, APC, PE, and MDA and the activities of SOD, POD, and CAT were significantly increased in the early cultivation and declined quickly at later stages. Finally, the algae-lysing mechanism of the bioactive metabolites of the bacteriaBrevibacillus laterosporusonOscillatoriahad been proposed.

Rapid Methods for Testing the Efficacy of Sterilization-Grade Filter Membranes

ABSTRACT The validation of sterilization-grade membranes is integral to ensuring the efficient and safe use of microfiltration systems. Here validation refers to the production of sterile filtrate for sterilizing-grade membranes under challenge test conditions. Current validation methods require 48 h of culture for results to become available, which creates time delays within the manufacturing process and quality control (QC) backlogs. This work compares four methods for the production of filter challenge test data, to the desired test sensitivity, within 24 h using bioluminescent and fluorescent recombinant strains of the test organism Brevundimonas diminuta. These methods should provide a way to implement more rapid QC test regimens for filters.

Enhancement of the infectivity ofFusobacterium necrophorumby other bacteria

SUMMARYNecrobacillosis is caused byFusobacterium necrophorum(FN), but other organisms are often present in the lesions. Their possible role was studied in experiments made with a virulent FN strain which, by itself, produced fatal necrobacillosis in mice provided that large doses ( > 106organisms, subcutaneously) were given. Mice were inoculated subcutaneously with FN suspended in sub-lethal doses (0·1 ml) of undiluted or diluted broth cultures of other bacteria. Undiluted culture of a strain ofEscherichia colireduced the infective dose of FX to < 10 organisms: in the necrobacillosis lesions that developed, fusobacteria greatly outnumberedE. coli. A heat-killed preparation or sterile filtrate ofE. coliculture had little if any effect on FN.Citrobacter freundiiand comparativelv small numbers ofCorynebacterium (Actinomyces) pyogenesproduced effects similar to that ofE. coli. An α-haemolvtic streptococcus.Pseudomonas aeruginosa.Bacteroides fragilisandFusobacterium nudeatumalso enhanced the infectivity of FN. though less strikingly thanE. coli. FN increased the persistence invivoof the α-haemolytic streptococcus andB. fragilis, and enabled the latter to multiply profusely.

PATHOGENICITY STUDIES WITH FUSARIUM POAE

An isolate of Fusarium poae (Pk.) Wr. from diseased pea plants at Ottawa caused complete wilting and death of several varieties and species of legumes within 72 hours of root inoculation. Tip-wilting and severe stunting symptoms were produced on inoculated barley, but no symptoms developed on wheat and oats. A sterile filtrate from a suspension of conidia and mycelium was phytotoxic when applied to the roots of peas and beans. The fungus was able to persist for more than 8 weeks in soil containing decomposing pea plants.

The mechanism of the training of Bact. lactis aerogenes to D-arabinose

Further experimental studies of the training of Bact. lactis aerogenes to utilize the pentose D-arabinose yield evidence against the presence in unadapted or partially adapted strains of the two distinct classes of normal and mutant cells. It has been found that the lag preceding colony formation by an unadapted strain on a solid medium is never less than the lag observed under comparable conditions in a liquid culture, but decreases continuously with the time of previous exposure to D-arabinose; that the presence of untrained cells does not inhibit the multiplication of rained cells; that the growth curve is not that characteristic of a simple mixture of normal cells and mutants; that at the end of the long lag phase the bacterial mass increases considerably before any detectable increase in cell number occurs, and that in favourable cases every cell present can become adapted. During the earliest stages of growth on D-arabinose no stably trained cells can be detected, all undergoing rapid reversion when grown in the presence of glucose under conditions where the selection of reverse mutants or of untrained cells could not be important. In mixtures of fully trained and normal cells there is no selective advantage in favour of the latter when prolonged subculture is made in glucose. The absence of reversion with fully trained cells cannot, therefore, be explained in this way (a matter relevant to a previous study). Co-operative effects result in a variation in lag according to the conditions of culture, and show themselves in the influence of sterile filtrate from grown cultures. The statistical variations in the length of the lag phase have been examined. It is concluded that no simple theory of mutation and selection will account for the observations, which, on the other hand, are adequately explained by a hypothesis of direct adaptation, i. e. the development on growth of an alternative metabolic route in the normal cell.

The mechanism of the training of Bact. lactis aerogenes to D-arabinose

Further experimental studies of the training of Bact. lactis aerogenes to utilize the pentose D-arabinose yield evidence against the presence in unadapted or partially adapted strains of the two distinct classes of normal and mutant cells. It has been found that the lag preceding colony formation by an unadapted strain on a solid medium is never less than the lag observed under comparable conditions in a liquid culture, but decreases continuously with the time of previous exposure to D-arabinose; that the presence of untrained cells does not inhibit the multiplication of trained cells; that the growth curve is not that characteristic of a simple mixture of normal cells and mutants; that at the end of the long lag phase the bacterial mass increases considerably before any detectable increase in cell number occurs, and that in favourable cases every cell present can become adapted. During the earliest stages of growth on D-arabinose no stably trained cells can be detected, all undergoing rapid reversion when grown in the presence of glucose under conditions where the selection of reverse mutants or of untrained cells could not be important. In mixtures of fully trained and normal cells there is no selective advantage in favour of the latter when prolonged subculture is made in glucose. The absence of reversion with fully trained cells can not, therefore, be explained in this way (a matter relevant to a previous study). Co-operative effects result in a variation in lag according to the conditions of culture, and show themselves in the influence of sterile filtrate from grown cultures. The statistical variations in the length of the lag phase have been examined. It is concluded that no simple theory of mutation and selection will account for the observations, which, on the other hand, are adequately explained by a hypothesis of direct adaptation, i. e. the development on growth of an alternative metabolic route in the normal cell.

The Toxicity of Sterile Filtrate from Parodontal Pockets

The local effect of the absorption of toxic material from pyorrhœa pockets on the hard and soft tissues around the teeth is well known. In this experiment an attempt was made to study the toxic effect on remote structures by injecting the sterile filtrate fresh from pyorrhœa pockets into various animals. The filtrate was obtained from patients with chronic pyorrhœa by removing the contents from parodontal pockets and passing them through a Seitz filter. The sterile filtrate obtained was then injected into cats, guinea-pigs, rabbits, and rats, in varying amounts. In the group of four cats, all showed fatty degeneration of the liver and two showed extreme fatty degeneration of the kidney tubules. Five guinea-pigs receiving one injection of ½ c.c. of filtrate showed no pathological change in the liver or kidney. One out of three guinea-pigs receiving two injections of filtrate showed fatty degeneration of the liver, while five out of six pigs receiving one injection of 1 c.c.,—i.e. double the quantity—showed definite fatty degeneration of the liver. One out of two rabbits showed similar changes and of six rats injected, all died in from five to seven days. The experiment suggests that substances are elaborated in parodontal pockets which are highly toxic and tend to injure the liver and kidney of animals in the process of their elimination. Such toxic material proved fatal in fifteen of the twenty-five animals injected.

FURTHER NOTES ON THE FILTRATION OF THE VIRUS OF VACCINIA

1. A very active filtrate can be obtained from vaccinia lesions by grinding up the fresh tissue with glass fragments, emulsifying in hormone broth, centrifuging the emulsion and filtering the supernatant fluid through a Berkefeld V filter. 2. The sterile filtrate so obtained has been shown by comparative titration on rabbits to have about one-sixteenth of the activity of the non-sterile emulsion used in human vaccination. 3. Centrifugation of such a filtrate shows a partial concentration of the virus in the lowermost layer. 4. The virus survives for a long time, if the filtrate is kept near the freezing point, and probably will survive indefinitely if kept frozen. The addition of glycerine is not necessary.

TISSUE-DIGESTING ENZYME (HISTASE) OF STREPTOCOCCI

1. An extracellular, proteolytic enzyme has been observed in more than 30 strains of beta type, aerobic and facultative hemolytic streptococci. 2. The enzyme is readily demonstrable in sterile filtrates of cooked meat cultures. 3. No gas or foul odor is produced. 4. It is partially inactivated by exposure to about 60°C. for 45 minutes or longer. 5. The enzyme manifests itself in cooked meat cultures after about 48 hours incubation at 37°C. The sterile filtrate from a 10 day old culture acts within 18 hours. 6. From 50 to 75 per cent of the meat in a tube of cooked meat medium may be digested in about 3 weeks at room temperature after 5 days initial growth at 37°C. 7. No correlation is found, in the cases studied, between hemolysis and proteolysis. 8. Streptococci not digesting beef tissue will not digest human tissue, and those which do digest beef tissue also digest human tissue. This conclusion applies only to the nine strains studied. 9. Ability to digest animal tissues does not necessarily imply ability to digest casein, coagulated beef serum or gelatin. 10. The disappearance of the meat from cooked meat cultures of hemolytic streptococci is quantitatively roughly paralleled by increase of formol-titrable substances in the fluid portion of the medium. 11. The enzyme resembles trypsin in its action. Streptococci from a variety of sources, bovine, human and otherwise have shown varying degrees of proteolytic activity. 12. The name histase is proposed for this enzyme.

THE ACTION OF PNEUMOCOCCUS ON BLOOD

The reduction of the oxygen capacity which occurs after incubating pneumococcus cultures with washed rabbit corpuscles is due to the formation of methemoglobin (or some derivative of hemoglobin with identical optical constants for three regions in the spectrum). The substance which induces the change is also present in the sterile filtrate of autolyzed cultures. By analogy we feel justified in concluding that the mechanism of the reduction of the oxygen capacity in human lobar pneumonia is at least in part of the same nature. To determine the frequency and intensity of the phenomenon in lobar pneumonia, and thereby to establish its clinical significance, is the next step and a problem upon which we are now engaged.