Mg Of Lysozyme Research Articles

High-level potentiation of lysostaphin anti-staphylococcal activity by lysozyme.

The purpose of this study was to determine whether lysostaphin would enhance its anti-staphylococcal efficacy in combination with lysozyme. Minimum inhibitory concentrations (MICs) of lysostaphin and lysozyme were separately determined for 41 strains belonging to 10 different species of human staphylococci. Lysozyme was virtually inactive and showed MICs of 15 mg/ml. On the contrary, all strains were susceptible to lysostaphin and showed MICs ranging from 2.5 to 60 micrograms/ml for the different Staphylococcus species. When the MIC of lysostaphin was determined in media containing submultiples of the MIC of lysozyme, the values obtained were much lower. The reduction of the lysostaphin MIC ranged from 16- to 200-fold in the different species tested. In Staphylococcus aureus, in particular, the combination of lysostaphin with 1.5 mg of lysozyme per ml reduced the MIC of lysostaphin by 25-fold. The activities of two combinations of the two enzymes were evaluated: one combination was expected to be active on S. aureus only, and the other combination was expected to inhibit all Staphylococcus strains. The first combination (0.5 micrograms of lysostaphin plus 0.5 mg of lysozyme per ml) was inhibitory to all of the 84 S. aureus strains tested, whereas 137 of 151 strains of other Staphylococcus species were unaffected. On the contrary, all of the 235 Staphylococcus strains tested were inhibited by the second combination (4 micrograms of lysostaphin plus 5 mg of lysozyme per ml). The possible mechanisms of lysostaphin potentiation by lysozyme are considered, and the potential use of a lysostaphin-lysozyme combination for topical therapy of staphylococcal infections resistant to other antibiotics is discussed.

Rapid immunochemical identification of enterococci with use of a centrifugal analyzer after a brief antigen-extraction step.

The precipitin reaction between antigens from enterococci and group-specific antisera was monitored turbidimetrically with a centrifugal analyzer. Linear standard curves (absorbance vs antigen concentration) were developed for studying optimal conditions for enzymic extraction of the antigen by use of an enzyme-containing filtrate from Streptomyces albus. This extraction was optimized by adding to the filtrate 1 mg of lysozyme per milliliter and incubating the bacteria for 40 min at 52 degrees C. The assay was adjusted so that detection of enterococci was indicated by positive absorbance changes near 0.1 A. When the concentration of bacteria from which the antigen was extracted was within a standardized range, non-enterococci produced no positive absorbance changes. Reproducibility of standard antigen and antisera solutions was excellent during three months. The procedure provides an objective, well-controlled means for identifying clinically important enterococci from pure cultures in about 1 h. In contrast, current methods involving biochemical media require as long as 48 h.

Selective antibacterial properties of lysozyme for oral microorganisms.

The antibacterial properties of lysozyme were investigated with oral microorganisms representing the seven serotypes (a through g) of Streptococcus mutans, Veillonella alcalescens, and the virulent (V) and avirulent (AV) strains of Actinomyces viscosus T14. Growth of bacteria in defined medium was monitored spectrophotometrically after the addition of various amounts (25 mug to 5 mg/ml) of enzyme. No growth inhibition of V. alcalescens was observed. Inhibition of A. viscosus T14(V) and A. viscosus T14(AV) occurred with 160 mug of lysozyme per ml. Of the S. mutans cultures tested, the serotype a and b strains were inhibited with as little as 25 mug of enzyme per ml, whereas e and f strains were most resistant to the bacteriostatic activity of lysozyme. The presence of dl-threonine or sucrose in growth medium did not significantly affect the results. A lysoplate assay was developed to rapidly survey the bacterial cultures for their susceptibility to the lytic ability of the enzyme. Lysis, as a measure of a zone of clearing in agarose plates, occurred for all microorganisms in the presence of lysozyme after the subsequent addition of NaCl or detergent. The bactericidal activity of lysozyme was determined on S. mutans BHT and S. mutans LM-7 by the pour plate technique. Preincubation of S. mutans LM-7 with as much as 1 mg of enzyme for 90 min did not affect viability or growth, whereas preincubation of S. mutans BHT with 1 mg of lysozyme resulted in no recoverable colony-forming units. An antigen containing extract of S. mutans LM-7 blocked the growth inhibitory property of lysozyme. Human lysozyme was a more effective antibacterial factor than hen egg white lysozyme. Total growth inhibition of S. mutans BHT was effected with 40 mug of human enzyme, and as little as 10 mug of human enzyme inhibited growth for greater than 20 h. The data presented indicate that different mechanisms may be responsible for the bacteriostatic, lytic, and bactericidal properties of the enzyme and that lysozyme is a selective but effective antibacterial factor for oral microorganisms.

Improved lysis of group N streptococci for isolation and rapid characterization of plasmid deoxyribonucleic acid

Procedures for effective cellular lysis and plasmid deoxyribonucleic acid (DNA) isolation from group N streptococci were developed. Cells were grown at 32 degrees C for 4 h in a modified Elliker broth containing 20 mM DL-threonine. After cellular digestion with 2 mg of lysozyme per ml for 7 min at 37 degrees C, 1% sodium dodecyl sulfate exposure resulted in complete and immediate lysis. Lactose (Lac) plasmid species in Streptococcus lactis C2 and S. cremoris B1 (30 and 37 megadaltons, respectively) were demonstrated upon examination of DNA from the cleared lysates by agarose gel electrophoresis. Increasing the lysozyme treatment to 20 min or more resulted in loss of the Lac plasmid, whereas other resident plasmids were unaffected and demonstrable in agarose gels. Diethylpyrocarbonate added before lysis prevented Lac plasmid loss in 20-min lysozyme-treated cells, but was not effective after 40 min of lysozyme treatment. The results suggested that endogenous nuclease activity during the lysozyme treatment period initiated Lac plasmid DNA loss. The development of an efficient lysis procedure for the group N streptococci allowed rapid identification and characterization of plasmid DNA by agarose gel electrophoresis. The plasmid composition of S. lactis C2 and S. cremoris B1, as determined by agarose gel electrophoresis, compared favorably to previous electron microscopic observations.

Active and Inactive States of Immunologic Unresponsiveness

Abstract Immunologic tolerance was induced in newborn mice by administration of 1, 5, 10, or 20 mg of lysozyme. These mice were subsequently challenged with a mixture of lysozyme and bovine serum albumin (BSA), or with a covalently coupled conjugate of lysozyme and succinylated BSA. When challenged at 6 weeks of age none of these mice responded to lysozyme when administered admixed with BSA. Of the mice challenged with the lysozyme-SBSA conjugate, those tolerized with 1 or 5 mg of lysozyme responded with antilysozyme antibodies but those tolerized with 10 or 20 mg of lysozyme did not. When challenged at 10 weeks of age, mice which had been tolerized with 20 mg of lysozyme mounted a low level antilysozyme response to both the mixture and the conjugate. Assessment of the response of these animals to BSA and SBSA revealed that the anti-SBSA responses of the conjugate-challenged animals were significantly suppressed whereas the anti-BSA response of the mixture-challenged animals was unaffected. Tolerance was also induced in adult mice, by the injection of 20, 50, or 100 mg of lysozyme. When challenged 12 days after cessation of the tolerizing regimen, all of these mice exhibited suppressed antilysozyme responses regardless of the challenge antigen, although the antilysozyme responses to the conjugate were slightly higher than to the mixture. No suppression of either anti-BSA or anti-SBSA responsiveness was seen in any of these adult tolerant animals. It was concluded that administration of low doses of tolerogen to neonatal mice results in only T cell tolerance whereas administration of higher doses of tolerogen also effects B cell responsiveness. Furthermore, neonatally induced tolerance appeared to involve some form of active suppression which did not appear to be a factor in the adult tolerized mice. This suppression was only demonstrable at the B cell level.

Serum and urinary lysozyme activities in patients with renal diseases.

Serum and urinary lysozyme values were assayed in 51 patients with chronic glomerulonephritis and 10 undergoing regular hemodialysis. Renal threshold for lysozyme was in the range from 15 to 20 μ/ml, but no correlation was obtained between serum and urinary lysozyme concentrations. Significant correlations were found between serum lysozyme activities and commonly employed parameters of renal dysfunctions, such as blood urea nitrogen level (r=0.77), serum creatinine level (r=0.86), and creatinine clearance (r=-0.80). The majority of patients with markedly diminished glomerular filtration rates smaller than 20 ml/min exhibited urinary excretion of more than 3 mg of lysozyme per day and elevated serum lysozyme, while in patients with mildly to moderately impaired renal function, serum and urine lysozyme concentrations were within normal limits. Therefore, positive lysozymuria and increased serum lysozyme activity may be taken as evidence of severe renal damage, whereas the lysozyme determination may not serve as sensitive indicators in mildly to moderately injured renal diseases.