Bacteriostatic action of progesterone on staphylococci and other microorganisms

Yotis, William (Loyola University, Hines, Ill.), and Ronald Stanke. Bacteriostatic action of progesterone on staphylococci and other microorganisms. J. Bacteriol. 92:1285-1289. 1966.-Progesterone has been examined in vitro for antibacterial activity against 10 microorganisms. Turbidimetric and manometric techniques were used to assay the antibacterial activity of progesterone. The organisms tested consisted of Staphylococcus aureus, S. epidermidis, Gaffkya tetragena, Bacillus subtilis, Listeria monocytogenes, Candida albicans, Escherichia coli, Aerobacter aerogenes, Salmonella paratyphi, and Proteus vulgaris. Antibacterial action was shown by progesterone only against the gram-positive microorganisms when they were grown in tryptic soy broth containing 10 to 20 mug of progesterone per ml. Pregnenolone, 4-pregnen-20beta-ol-3-one, and 5alpha-pregnane also possessed antistaphylococcal properties, whereas pregnanolone, pregnandione, 11alpha-hydroxyprogesterone, and 17alpha-hydroxyprogesterone did not. The bacteriostatic action of progesterone on staphylococci was exerted primarily during the first 8 hr of incubation, and it was reduced in the presence of oxygen. In the presence of 20 mug of progesterone per ml, there was significant reduction in the oxidation by resting staphylococcal suspensions or utilization by staphylococci of pyruvate as an energy source during growth.     

Effect of Inoculum Size on In Vitro Susceptibility Testing with Lincomycin

There is disagreement in the literature as to whether lincomycin is primarily a bacteriostatic or a bactericidal agent against gram-positive cocci and also regarding the levels of activity of this agent against susceptible microorganisms. These questions were examined in a study of the effect of inoculum size on the results of tube dilution susceptibility determinations with lincomycin against 49 clinical isolates of Staphylococcus aureus and 25 strains of streptococci and pneumococci. Lincomycin was both highly active and bactericidal when tested against 40 strains of S. aureus with inocula containing a maximum of 10(4) cells per ml [median minimal inhibitory concentration (MIC), 0.78 mug/ml; median minimal bactericidal concentration (MBC), 1.56 mug/ml]. With inocula of 10(5) cells per ml, lincomycin was primarily bacteriostatic (median MIC, 1.56 mug/ml; median MBC, 12.5 mug/ml). There were further decreases in inhibitory levels and significant losses of bactericidal activity when inocula containing more than 10(7) cells were tested (median MIC, 3.13 mug/ml; median MBC > 100 mug/ml). Similar measurements with streptococci and pneumococci revealed a lesser effect of inoculum size. The mean MBC value for alpha-hemolytic streptococci increased from 0.40 to 1.05 mug/ml with an increase in inocula from 10(4) to 10(6) cells per ml, but without a marked increase in MIC values. Similar results were obtained for beta-hemolytic streptococci and pneumococci.     

Cephapirin: In Vitro Antibacterial Spectrum

Cephapirin, a new semisynthetic cephalosporin derivative, was found to have an antibacterial spectrum similar to that of cephalothin. Staphylococcus aureus was inhibited by cephapirin concentrations of 0.09 to 12.5 mug/ml. S. epidermidis, S. viridans, S. pyogenes, and Diplococcus pneumonia isolates were inhibited by less than 1 mug/ml. The Enterococcus required a concentration of 25 mug of antibiotic per ml for inhibition. Approximately 65% of Escherichia coli, and all Klebsiella, indole-negative Proteus, and Salmonella strains tested were inhibited by the drug. Serratia, Pseudomonas, indole-positive Proteus, and Erwinia strains were highly resistant. Inoculum size was not an important factor in determining the level of sensitivity of S. aureus to cephapirin. The antibiotic does not appear to be significantly bound to serum protein. In vitro development of resistance to the drug was demonstrated with two isolates of S. aureus.     

Responses of Staphylococci to Androgens

The reported regulatory activities of hormones on mammalian cells suggest that a hormonal effect may be of importance in the host-parasite relationship of staphylococci. Male 6-month-old rabbits of similar genetic constitution were given subcutaneously 20 mug of androgens in saline containing 1% ethyl alcohol. Control rabbits received 1% ethyl alcohol in saline. At 5 to 10 min after administration of androgens, the rabbits were bled by cardiac puncture, and the serum was separated and incubated with standardized suspensions of Staphylococcus aureus serotypes I to XIII. It was found that S. aureus grew more luxuriantly in the sera of the control rabbits than in the sera of androgen-treated animals. With tryptic soy broth as a culture medium, a concentration 150- to 300-fold (30 to 40 mug/ml) higher than that achieved in the blood of an androgen-treated rabbit was required to yield an equivalent effect. In addition, the androgen-staphylococcal interaction has been studied with regard to experimentally induced furunculosis, the uptake of androgens by staphylococci and steroid molecular structure and antimicrobial activity. The data indicate that androgens may play a role in protection against staphylococcal infection.     

In Vitro Effects of Some Chemotherapeutic Agents on Mycobacteria

Examination of the bactericidal and bacteriostatic effects of nine antibiotics on atypical mycobacteria revealed that streptovaricin complex and streptovaricin C exerted bactericidal effects on several strains in concentrations lower than 1.0 mug/ml. An exposure to the drug for 48 hr at 37 C was necessary to effect a complete inactivation of more than 99.9% of the exposed microorganisms. The appearance of strepto-varicin-resistant mutants was observed. However, these mutants were unstable, and reversion to streptovaricin susceptibility occurred. Celesticetin salicylate, added in a concentration of 100.0 mug/ml to the medium of Olitzki and Gershon inoculated with Mycobacterium leprae, effected a complete change of the uniformly stained mycobacteria to bipolarity, which indicates the devitalization of this microorganism.     

Antimicrobial properties of testosterone and its intermediates

Testosterone, epiandrosterone, dehydroisoandrosterone and androsterone have been examinedin vitro for antibacterial activity againstStaphylococcus aureus, S. epidermidis, Listeria monocytogenes, Streptococcus faecalis, Escherichia coli, Alcaligenes faecalis, andSalmonella paratyphy. Turbidimetric and manometric techniques were used to assay the antibacterial activity of the above androgens. Antibacterial action was shown by epiandrosterone and dehydroisoandrosterone only against the gram-positive microorganisms when they were grown in tryptic soy broth containing 10 to 20µg of hormone per ml.The oxidation of pyruvate in the presence of 20µg/ml of epiandrosterone and dehydroisoandrosterone gave lower Qo₂ than in the absence of the hormones. In the presence of 20µg per ml of culture medium and under anaerobic conditions, testosterone and androsterone also possessed antistaphylococcal activity. In vivo tests indicate that a dose ofS. aureus twice as large was required to produce skin lesions in 50% of 6-month-old male rabbits given subcutaneously a total of 30µg of testosterone over a period of 2 weeks than in control animals. Furthermore, the size of erythema surrounding the site of injection of staphylococci or staphylococcal alpha toxin was significantly smaller in the testosterone-treated rabbits than in the control. Thus, androgens may influence resistance to furunculosis.     

Combined Effects of Diphenyliodonium Chloride, Pine Oils, and Mustard Oil Soaps on Certain Microorganisms

Bactericidal and bacteriostatic activities of an emulsion containing 10.0% (v/v) terpineol, 0.5% (w/v) diphenyliodonium chloride, 11.0% (v/v) ethyl alcohol, and 5.62% saponified mustard oil were tested against a number of different types of organisms. The bactericidal concentration for Salmonella typhosa was 1:400. In the presence of 5.0% horse serum, it increased to 1:250. The bacteriostatic concentration varied from organism to organism; Escherichia coli and Staphylococcus aureus required 4,000 mug/ml for complete bacteriostasis, whereas Corynebacterium diphtheriae, Salmonella paratyphi-A, and Shigella required only 2,000 mug/ml for complete inhibition. A 4.0% concentration of the emulsion killed the spores of Bacillus subtilis within 6 hr.     

Biological Characterization of Prasinomycin, a Phosphorus-containing Antibiotic

Prasinomycin, a new antibiotic from the green spore streptomycete, Streptomyces prasinus, primarily inhibits the growth of gram-positive microorganisms. Like penicillin, it is effective only against growing cells. Though primarily bacteriostatic at levels about the minimal inhibitory concentration, it is bactericidal at higher levels. Neither synergism nor antagonism could be demonstrated for prasinomycin with a variety of other antibiotics. It is highly active upon subcutaneous administration to mice infected with Staphylococcus aureus, Streptococcus pyogenes C203, or Diplococcus pneumoniae. Prasinomycin has a unique prophylactic action whereby one dose protects mice against experimental infections for as long as 2 months. It is more effective against S. aureus infections in mice when administered subcutaneously 20 hr prior to infection than when given in divided doses 1 hr before and 4 hr after infection.     

Polymyxin-Coagulase-Mannitol-Agar: I. A Selective Isolation Medium for Coagulase-Positive Staphylococci1

A selective, differential plating medium was developed for the isolation and identification of coagulase-positive and mannitol-fermenting staphylococci. Coagulase produced by growing Staphylococcus aureus caused an opaque zone of fibrin to form around each colony. Several strains of S. aureus produced a visible coagulase reaction by 8 hr, and all strains gave a positive reaction before 12 hr. Mannitol fermentation was usually observed between 12 and 36 hr. Rabbit serum was filtered through Sephadex G-100 to obtain plasmin- and plasminogen-free coagulase-reacting factor (CRF). False-negative reactions, caused by staphylokinase and staphylococcal Müller factor action on plasminogen, were eliminated when this CRF was used. False-positive reactions by lipolytic, coagulase-negative staphylococci were reduced, since gel filtration removed the serum lipoprotein which served as a primary source of opacity. The addition of 75 mug of polymyxin B per ml selectively retarded the growth of S. epidermidis and minimized false-positive reactions caused by citrate-utilizing gram-negative rods. The preparation, characteristics, and use of the medium are presented.     

Action of Diethylstilbestrol on Staphylococci: Further Observations on the Effect of Resting Cells

Diethylstilbestrol (DS) has been shown to be active against staphylococci and other gram-positive bacteria but not against gram-negative microorganisms. The present study extends these findings. Standardized suspensions of (14)C-labeled Staphylococcus aureus serotypes III and IV and Shigella flexneri were prepared and exposed to pharmacological concentrations of DS (1 to 20 mug/ml) under diverse environmental conditions; the cells were removed by membrane filtration and the presence of radioactive substances in release to the supernatant fraction was followed by standard radioisotopic techniques. Controls were exposed similarly to the hormone vehicle alone (buffer containing 2% ethyl alcohol). DS at bactericidal concentrations above 6 mug/ml caused significant leakage of cellular radioactivity of S. aureus labeled with (14)C-glucose and (14)C-glutamic acid within 1 to 4 hr after exposure to DS. Maximum leakage of radioactivity occurred under anaerobic conditions at 37 C. Absorption studies of (14)C-labeled DS indicated that the affinity of S. flexneri for DS is markedly less than that of S. aureus. This might be one reason for the resistance of gram-negative bacteria to DS.     

Production of Enterotoxin A in Milk

Enterotoxin A production in milk was studied by use of variables of milk quality, initial numbers of enterotoxigenic staphylococci, incubation temperature, and time. In both raw and pasteurized milks having a low total viable count, enterotoxin was detected in minimal incubation times of 6 to 9 hr at 35 C, 9 to 12 hr at 30 C, 18 hr at 25 C, and 36 hr at 20 C, after inoculation with 10(6)Staphylococcus aureus cells per ml. When similar milks were inoculated with 10(4)S. aureus cells per ml, enterotoxin was detected in 12 hr at 35 C, 18 hr at 30 C, 24 to 36 hr at 25 C, and 48 to 96 hr at 20 C. In high-count raw milk, enterotoxin was detected only in samples inoculated with 10(6)S. aureus cells per ml and incubated at 35 C. Generally, a concentration of 5 x 10(7)S. aureus cells per ml of milk was reached before enterotoxin A was detected.     

The antibacterial activity against MRSA strains and other bacteria of a <500Da fraction from maggot excretions/secretions of Lucilia sericata (Diptera: Calliphoridae)

The application of Lucilia sericata larvae to chronic, infected wounds results in the rapid elimination of infecting microorganisms, including MRSA. Previously, we demonstrated in vitro antibacterial activity of native excretions/secretions (nES) from L. sericata and partially purified two low mass antibacterial compounds with masses of 0.5-10kDa and <500Da. The present study reports the antibacterial effects of the <500Da fraction (ES<500) on the growth and morphology of a range of bacteria, including 12 MRSA strains. Distinct morphological changes were observed in Bacillus cereus and Escherichia coli following exposure to ES<500. Flow cytometry and confocal microscopy analyses, in conjunction with turbidometric and CFU assays, revealed bacteriostatic activity of nES against S. aureus and E. coli. ES<500 also demonstrated bacteriostatic activity against S. aureus, however, bactericidal activity and the induction of a viable but non-culturable state were observed with ES<500-treated E. coli.     

The effect of difluorodichloromethane (FC 12) on isolated rat and rabbit heart]

We have tried to determine if dichlorodifluoromethane (F.C. 12) might have an effect in vitro on the isolated rat and rabbit hearts. The direct action of F.C. 12 on the heart in vitro is similar in both rats and rabbits. It occurs at doses such as 20 +/- 10 mug/ml (rat) and 35 +/- 5 mug/ml (rabbit). F.C. 12 depresses the strength of the myocardial contractions; the effect is reversible at low concentration (less than 60 mug/ml) but irreversible at high concentration (greater than or equal to 120 mug/ml). It also causes a slight bradycardia, but no significant effect on the basal tonus. At high concentration, F.C. 12 may produce arrhythmia : this action occurs more readily in the rat heart than in the rabbit heart.     

In vitro inhibitory effect of cranberry (Vaccinium macrocarpom Ait.) juice on pathogenic microorganisms

The purpose of this study was to determine the inhibitory effects of cranberry juice on pathogenic microorganisms. The microorganisms analyzed were Escherichia coli from patients with urinary infections, Salmonella spp., Listeria monocytogenes, Pseudomonas aeruginosa, and Staphylococcus aureus. The disc method was used to determine the sensitivity of bacteria to cranberry juice (CJ, both concentrated and diluted). A lawn of 10(6) cfu/ml was grown on agar surfaces in Petri dishes and on Whatman discs that had been previously saturated with CJ and CJ : water 1 : 1 to 1 : 50 juice solutions had been placed on the discs, which were cultured and incubated. The results indicated that S. aureus was more susceptible to cranberry juice inhibition than the other microorganisms. L. monocytogenes was the most resistant to the inhibitory action of cranberry juice, showing a significant difference from the inhibition of P. aeruginosa, uropathogenic E. coli, Salmonella spp., and S. aureus. This study also demonstrated that the inhibitory activity of cranberry juice for E. coli took place up to a dilution of 1 : 20.     

Evidence for a Multiplicity of Capsular Types Among Staphylococcus aureus Strains

The Smith diffuse variant and the wound mucoid strain of Staphylococcus aureus were shown to exhibit serologically distinct capsules. The Welwood and K-6 strains of S. aureus were tested to determine their capsular types. Both Welwood and K-6 were found to be representative of the Smith capsular type. An additional 13 isolates of S. aureus from mice were tested. Gel double-diffusion tests and immunoelectrophoresis of staphylococcal antigens disclosed the possible existence of at least two additional capsular types. Passive hemagglutination tests carried out with cells sensitized with 1 mg of antigen per ml showed a multiplicity of cross-reacting antigens. However, cells sensitized either with 0.1 or 0.05 mg of antigen per ml and reacted with antisera absorbed with 10 or 1 mug/ml showed the presence of a specific antigen in each strain of S. aureus. Corroborative evidence for a multiplicity of capsular types was obtained by the specific capsular reaction. At least four capsular types of S. aureus were found. The prototypic strains for these antigens are the RLM or wound strain, the Smith diffuse strain, and mouse strains designated 36T and 43R. We propose to designate these types 1, 2, 3, and 4, respectively.     

In Vitro Antimicrobial Activity and Human Pharmacology of Cephaloglycin

Serum and urine concentrations of cephaloglycin (an orally absorbed derivative of cephalosporin C) were determined in normal volunteers and in patients. The in vitro activity of cephaloglycin was also studied. All strains of group A streptococci (Streptococcus pyogenes) and Diplococcus pneumoniae were inhibited by 0.4 mug of cephaloglycin per ml. Eighty per cent of the Staphylococcus aureus strains and about 50% of the Escherichia coli and Proteus mirabilis strains were inhibited by 1.6 mug of cephaloglycin per ml. Klebsiella-Aerobacter species were more resistant to cephaloglycin and 12.5 mug per ml was required to inhibit 70% of these strains. When single doses of 250, 500, or 1,000 mg of cephaloglycin were administered to fasting volunteers, a peak serum concentration of at least 0.5 mug per ml was achieved. A full breakfast did not interfere with absorption of cephaloglycin. Probenecid enhanced both the peak serum concentration and the duration of antibiotic activity in the serum. Serum concentrations of cephaloglycin were even higher in patients who were receiving repeated doses. The peak serum concentrations of cephaloglycin in all volunteers and patients were adequate to inhibit all strains of group A streptococci and D. pneumoniae. Many of the peak serum concentrations were adequate to inhibit some strains of S. aureus, E. coli, and P. mirabilis. Urine levels of cephaloglycin were high enough in all volunteers and patients to inhibit more than 90% of the E. coli and P. mirabilis strains and over 70% of the strains of Klebsiella-Aerobacter.     

The killing effect of cryptolepine on Staphylococcus aureus

AIM: This study was undertaken to further examine the antimicrobial actions of the alkaloid cryptolepine. METHODS AND RESULTS: The minimum inhibitory concentration (MIC) of cryptolepine against Staphylococcus aureus was determined using the broth dilution method. Time-kill kinetics and scanning electron microscopy (SEM) techniques were employed to monitor the survival characteristics and the changes in morphologies respectively of staphylococci in the presence of cryptolepine. A notable antistaphylococcal activity was recorded for cryptolepine (MIC against S. aureus NCTC 10788=5 microg ml(-1)). Cryptolepine appears to have a lytic effect on S. aureus as seen in SEM photomicrographs following 3, 6 or 24 h treatment with 4X MIC, i.e. 20 microg ml(-1) of cryptolepine. The surface morphological appearance of the staphylococcal cells was also altered. The lytic effect appeared to coincide with low viable counts recorded in survival curves following treatment with cryptolepine. SIGNIFICANCE AND IMPACT OF THE STUDY: These findings demonstrate that lysis occurs when susceptible organisms are exposed to cryptolepine.     

Convenient Assay for Staphylococcal Nuclease by the Metachromatic Well-Agar-Diffusion Technique

The metachromatic agar-diffusion (MAD) microslide technique was adapted for quantitative assay for staphylococcal thermonuclease in heterogeneous systems, such as milk and broth. When an enzyme-containing solution was placed in a well cut in the agar, a bright pink halo was obtained. The diameter of the pink zone of hydrolysis was related to time and temperature of incubation and to nuclease concentration. Concentrations of nuclease as low as 0.005 mug/ml and as high as 2.0 mug/ml were conveniently determined after 3 hr at 37 C.     

Effect of prostaglandins on cell function and multiplication of parainfluenza 3 viruses in WISH cells.

Cytotoxic effect of prostaglandins E2 and F2alpha on cells grown in vitro and the influence of these compounds on multiplication of myxovirus parainfluenza 3 were investigated. The prostaglandins were added to culture medium (0-01-10 mug/ml) 24 hr before virus infection, or for 2 and 48 hr after inoculation with viruses. WISH cells and monkey kidney cell cultures were used. No direct cytotoxic effect of prostaglandins at concentrations 0-01-1 mug/ml was found (viability, supravital staining, phase-contrast system, Nitro-BT reduction and succinic dehydrogenase tests), whereas the concentration of 10 mug/ml within 48 hr led to reduction and succinic dehydrogenase tests), whereas the concentration of 10 mug/ml within 48 hr led to partial injury of the cell population with symptoms of damage to mitochondria. Prostaglandins E2 and F2alpha inhibited multiplication of parainfluenza 3 virus at concentrations 0-1-10 mug/ml. The inhibitory effect was most pronounced if prostaglandins were added to medium for the whole period of virus multiplication i.e. for 48 hr but little or no effect was found if they were added prior to inoculation or for 2 hr after it. Inhibitory effect of prostaglandins on replication phase of viruses is suggested.