Injury to Staphylococcus aureus during sausage fermentation.

Staphylococcus aureus 196E added to a beef sausage containing starter culture and 0.5 to 2.0% glucose and incubated at 35 degrees C was unable to grow when plated on tryptic soy agar (TSA) containing 7.5% NaCl. The injury, presumed to be due to the lactic acid produced during fermentation, was more pronounced at the lower concentrations of glucose (and lower acid levels). In the absence of glucose and/or starter culture, no injury was observed. When sausages containing S. aureus injured by fermentation at 35 degrees C were incubated at 5 degrees C, the counts on TSA (measures both injured and uninjured cells) and TSA containing 7.5% NaCl (measures uninjured cells only) remained constant; however, upon reincubation of the cold-stored sausage at 35 degrees C, the staphylococcus counts on TSA and TSA containing 7.5% NaCl and were similar to the counts of S. aureus present in fermenting sausages that had never been subjected to 5 degrees C. The demonstration of acid injury indicated that the injury phenomenon must be considered when determining numbers of viable S. aureus in fermented sausages.     

The survival of Staphylococcus aureus during the fermentation and storage of yoghurt

The effect of yoghurt culture R _x on the survival of Staphylococcus aureus CCM 5984 added to milk in various concentrations was observed during the fermentation and storage of yoghurt. The end of the fermentation process (3.5 h) was only accompanied by a slight reduction. During the storage of yoghurt at 4°C a 1-2 log reduction was observed. No Staph. aureus was detected in yoghurt produced from milk contaminated by 10³ Staph. aureus cells 1^-1 after 48 h of cold storage. When a concentration of 10² Staph. aureus cells was used for milk contamination, the pathogen was not recovered from yoghurt during the fermentation and storage. The fermentation and storage of yoghurt was accompanied by increases in lactic acid and titrimetric acidity, as well as by a decrease in pH value.     

Enterotoxin B formation by fermentation mutants of Staphylococcus aureus.

An appreciable fraction of carbohydrate-negative (car) mutants of Staphylococcus aureus strains ATCC 14458, 778, and S-6 exhibit increased enterotoxin B (SEB) production. In addition, some lac and mtl mutants of these strains also display enhanced SEB formation. All such mutants appear to be point mutations. Mutagen-induced reversions of high SEB producing car, mtl, or lac mutants yield varying amounts of SEB and some clones seem to be restored to the characteristics of the parent type. A few sequentially isolated lac, mtl double mutants of strain 778 elaborate much more or much less SEB than either the lac or the mtl single mutants.     

Destruction of Staphylococcus aureus during frankfurter processing.

We studied the thermal resistance of Staphylococcus aureus during frankfurter processing in respect to whether staphylococci are killed by the heating step of the process and whether heat injury interferes with the quantitative estimation of the survivors. With S. aureus 198E, heat injury could be demonstrated only when large numbers of cells (10(8)/g) were present and at a product temperature of 140 degrees F (60 degrees C). On tryptic soy agar and tryptic soy agar plus 7% NaCl media, at temperatures less than 140 degrees F, the counts were virtually identical; above 140 degrees F, the counts converged, with the organisms dying so rapidly that heat injury was not demonstrable. Heat injury was thus judged not to interfere with the quantitative estimation of staphylococci surviving the normal commercial heating given frankfurters. By using a combination of direct plating on tryptic soy agar and a most-probable-number technique, we detected no viable cells (less than 0.3/g) of several strains of S. aureus in frankfurters heated to 160 degrees F (71.1 degrees C). This temperature is compatible with the normal final temperature to which federally inspected processors heat their frankfurters and with the temperature needed to destroy salmonellae.     

Binding of elastin to Staphylococcus aureus.

Many pathogenic bacteria specifically bind to components of the extracellular matrix. In this study, we report the specific association of Staphylococcus aureus with elastin, a major structural component of elastic tissue. Competition assays in which the binding of radiolabeled tropoelastin was inhibited by excess unlabeled elastin peptides, but not by other proteins, established the specificity of the interaction. Kinetic studies showed that tropoelastin binding to the bacteria was rapid and saturable. Scatchard analysis of the equilibrium binding data indicated the presence of a single class of high affinity binding sites (KD approximately 4-7 nM) with approximately 1000 sites per organism. Protease susceptibility suggested that the elastin binding moiety on S. aureus was a protein, which was confirmed by the isolation of a 25-kDa elastin-binding protein from S. aureus extracts through affinity chromatography. Using a truncated form of tropoelastin, the bacterial binding domain on elastin was mapped to a 30-kDa fragment at the amino end of the molecule. Although the precise amino acid sequence recognized by the staphylococcal elastin receptor has not been characterized, it is clearly different from the region of tropoelastin that specifies binding to mammalian elastin receptors.     

Ribosome assembly during recovery of heat-injured Staphylococcus aureus cells.

The process of ribosome formation during repair of sublethal heat injury was examined in Staphylococcus aureus. Sublethal heating of this organism results in the degradation of the 30S ribosomal subunit and alteration of the 50S subunit. Cells recovering from sublethal injury were examined for changes with time in the sedimentation and electrophoretic properties of ribonucleoprotein particles and ribonucleic acid, respectively. When cells were allowed to recover in [3H]uridine, the label could be followed into ribonucleic acid species that coelectrophoresed with 23S and 16S ribonucleic acid. Three ribonucleoprotein particles (49S, 36S, and 30S) were isolated from repairing cells by sedimentation through sucrose gradients. Polyacrylamide gel electrophoresis showed that the 49S particle contained 23S ribonucleic acid, the 36S particle contained both 23S ribonucleic acid and 16S precursor and mature ribonucleic acid, and the 30S particle contained 16S and precursor 16S ribonucleic acid. Particles with similar sedimentation properties were found in unheated cells.     

Epidemiology of Staphylococcus aureus during space flight

Abstract Staphylococcus aureus was isolated over 2 years from Space Shuttle mission crewmembers to determine dissemination and retention of bacteria. Samples before and after each mission were from nasal, throat, urine, and feces and from air and surface sampling of the Space Shuttle. DNA fingerprinting of samples by digestion of DNA with Sma I restriction endonuclease followed by pulsed-field gel electrophoresis showed S. aureus from each crewmember had a unique fingerprint and usually only one strain was carried by an individual. There was only one instance of transfer between crewmembers. Strains from interior surfaces after flight matched those of crewmembers, suggesting microbial fingerprinting may have forensic application.     

Staphylococcus aureus surface protein SasG contributes to intercellular autoaggregation of Staphylococcus aureus

Staphylococcus aureus surface protein G (SasG) is one of cell surface proteins with cell-wall sorting motif. The sasG mutant showed significantly reduced cell aggregation and biofilm formation. SasG is comprised of variable A domain and multiple tandem repeats of B domain, native-PAGE and in vitro formaldehyde cross-linking experiments revealed that the recombinant protein of the A domain showed homo-oligomerization as an octamer, but B domain did not. This study shows that SasG-A domain contributes to intercellular autoaggregation by homo-oligomerization, and that may facilitate the adherence to host-tissues in the infection of S. aureus.     

Alpha-toxin of Staphylococcus aureus.

Alpha-toxin, the major cytotoxic agent elaborated by Staphylococcus aureus, was the first bacterial exotoxin to be identified as a pore former. The protein is secreted as a single-chain, water-soluble molecule of Mr 33,000. At low concentrations (less than 100 nM), the toxin binds to as yet unidentified, high-affinity acceptor sites that have been detected on a variety of cells including rabbit erythrocytes, human platelets, monocytes and endothelial cells. At high concentrations, the toxin additionally binds via nonspecific absorption to lipid bilayers; it can thus damage both cells lacking significant numbers of the acceptor and protein-free artificial lipid bilayers. Membrane damage occurs in both cases after membrane-bound toxin molecules collide via lateral diffusion to form ring-structured hexamers. The latter insert spontaneously into the lipid bilayer to form discrete transmembrane pores of effective diameter 1 to 2 nm. A hypothetical model is advanced in which the pore is lined by amphiphilic beta-sheets, one surface of which interacts with lipids whereas the other repels apolar membrane constitutents to force open an aqueous passage. The detrimental effects of alpha-toxin are due not only to the death of susceptible targets, but also to the presence of secondary cellular reactions that can be triggered via Ca2+ influx through the pores. Well-studied phenomena include the stimulation of arachidonic acid metabolism, triggering of granule exocytosis, and contractile dysfunction. Such processes cause profound long-range disturbances such as development of pulmonary edema and promotion of blood coagulation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Staphylococcus aureus Panton-Valentine Leukocidin Contributes to Inflammation and Muscle Tissue Injury

Community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) threatens public health worldwide, and epidemiologic data suggest that the Panton-Valentine Leukocidin (PVL) expressed by most CA-MRSA strains could contribute to severe human infections, particularly in young and immunocompetent hosts. PVL is proposed to induce cytolysis or apoptosis of phagocytes. However, recent comparisons of isogenic CA-MRSA strains with or without PVL have revealed no differences in human PMN cytolytic activity. Furthermore, many of the mouse studies performed to date have failed to demonstrate a virulence role for PVL, thereby provoking the question: does PVL have a mechanistic role in human infection? In this report, we evaluated the contribution of PVL to severe skin and soft tissue infection. We generated PVL mutants in CA-MRSA strains isolated from patients with necrotizing fasciitis and used these tools to evaluate the pathogenic role of PVL in vivo. In a model of necrotizing soft tissue infection, we found PVL caused significant damage of muscle but not the skin. Muscle injury was linked to induction of pro-inflammatory chemokines KC, MIP-2, and RANTES, and recruitment of neutrophils. Tissue damage was most prominent in young mice and in those strains of mice that more effectively cleared S. aureus, and was not significant in older mice and mouse strains that had a more limited immune response to the pathogen. PVL mediated injury could be blocked by pretreatment with anti-PVL antibodies. Our data provide new insights into CA-MRSA pathogenesis, epidemiology and therapeutics. PVL could contribute to the increased incidence of myositis in CA-MRSA infection, and the toxin could mediate tissue injury by mechanisms other than direct killing of phagocytes.     

Monitoring of Staphylococcus xylosus DSM 20266 added as starter during fermentation and ripening of soppressata molisana, a typical Italian sausage

AIMS: "Soppressata molisana", a fermented sausage produced in southern Italy, is commonly obtained without starter addition. However, the use of starter cultures is more and more recommended in meat fermentation processes in order to guarantee stable production performance. In this study, the survival of the Staphylococcus xylosus DSM 20266 was evaluated during the ripening of "soppressata molisana" fermented sausage. METHODS AND RESULTS: The fastest method of RAPD-PCR was employed for discrimination of the added strain from those naturally present during the ripening of the "soppressata molisana". The results obtained were confirmed by analysis of the DNA macrorestriction profile by PFGE. The electrophoretic pattern of bacterial total proteins was also studied, but clear differences between the different strains could not be detected. CONCLUSIONS: The RAPD technique was a valid tool for monitoring Staph. xylosus DSM 20266 in "sopressata molisana". SIGNIFICANCE AND IMPACT OF THE STUDY: This study highlights the possibility of monitoring the presence of Staph. xylosus strains during the ripening of fermented sausages by a reliable and repeatable technique such as RAPD.     

Staphylococcal polysaccharide products inhibitory to Staphylococcus aureus bacteriophage.

Four products derived from Staphylococcus aureus cultures were partially purified and tested for inhibitory activity against staphylococcal phage. Phage inhibition, a specific stable phenomenon, was concentration dependent. All inhibitory products contained carbohydrate and amino acids, the most active (phage 73 lysate product) having a high carbohydrate content. Galactose, glucosamine, five or six amino acids, and possibly 3-O-methylglucose and a uronic acid were found as components in all active preparations. However, the exact nature of the active material remains undetermined.     

Mechanism of Thermal Injury in Staphylococcus aureus: I. Relationship Between Viability and Leakage

The viability of and leakage from suspensions of Staphylococcus aureus stored in the temperature range 3 to 60 C were determined. There was a direct relationship, up to 50 C, between death and leakage, both of which increased with increasing temperature. At temperatures above 50 C, there was a different pattern of response, the total leakage being less at 60 C than at 50 C, although there was greater membrane damage at the higher temperature. Sucrose, 1 m, almost completely prevented leakage at 37 and 50 C but not at 60 C.     

Regeneration of Ribosomes and Ribosomal Ribonucleic Acid During Repair of Thermal Injury to Staphylococcus aureus

Heating Staphylococcus aureus MF31 at 55 C for 15 min renders the organisms unable to reproduce on agar containing 7.5% NaCl (1). The heated organisms exhibited an extended lag period during which the organisms regained their ability to grow on the 7.5% NaCl-agar. Inhibitor and antibiotic data indicated that protein synthesis is not involved in this recovery process but nucleic acid synthesis is suggested (3). The data presented here further substantiate the noninvolvement of protein synthesis during recovery and further demonstrate the site of the thermally induced nucleic acid lesion. Methylated albumin kieselguhr column analysis showed the lesion site to be the ribosomal ribonucleic acid (rRNA). The rRNA is resynthesized during the extended lag period. Sucrose gradient analysis demonstrated that a ribosomal peak was undetectable subsequent to the thermal treatment, but this peak was regenerated during the recovery period.