Bovine whey proteins inhibit the interaction of Staphylococcus aureus and bacteriophage K

Abstract

Aims: To understand the potential use of bacteriophage K to treat bovine Staphylococcus aureus mastitis, we studied the role of whey proteins in the inhibition of the phage–pathogen interaction in vitro. Methods and Results: The interaction of bacteriophage K and S. aureus strain Newbould 305 was studied in raw bovine whey and serum. Incubation of S. aureus with phage in whey showed that the bacteria are more resistant to phage lysis when grown in whey and also bovine serum. Whey collected from 23 animals showed a wide variation in the level of phage‐binding inhibition. The role of the protein component of milk whey in this inhibition was established; treatment of the whey by heat, proteases and ultrafiltration removed the inhibitory activity. Brief exposure of S. aureus cells to whey, followed by resuspension in broth, also reduced phage binding. Microscopy showed the adhesion of extracellular material to the S. aureus cell surface following exposure to whey. Chromatographic fractionation of the whey demonstrated that the inhibitory proteins were present in the high molecular weight fraction. Conclusions: The adsorption of whey proteins to the S. aureus cell surface appeared to inhibit phage attachment and thereby hindered lysis. The inhibitory whey proteins are of high molecular weight in their native form and may sterically block phage attachment sites on the cell surface. Significance and Impact of the Study: These findings have implications for any future use of phage therapy in the treatment of mastitis, and other diseases, caused by S. aureus. This pathogen is predicted to be much more resistant to phage treatment in vivo than would be expected from in vitro broth culture experiments.

Efficacy and Pharmacokinetics of Bacteriophage Therapy in Treatment of Subclinical Staphylococcus aureus Mastitis in Lactating Dairy Cattle

“…Additionally, a reduction in the binding rate of phage K to S. aureus which is independent of phage inactivation has been observed in milk or milk whey (14,24). There is some evidence that this rate reduction is mediated by whey proteins which attach to the S. aureus cell surface and inhibit phage binding (14).…”

Bacteriophage biocontrol of foodborne pathogens

“…In curd a reduction of 4.64 log CFU per g was obtained compared with control Bueno et al 2012 Combination of HPP (high pressure processing) and phage resulted in S. aureus elimination in pasteurized milk within the 48 h regardless of the initial contamination level (1×10 6 or 1×10 4 CFU per mL) Tabla et al 2012 Phage cocktail (Φ88 and Φ35) along with nisin application decreased S. aureus by 1 log unit more than phage or nisin applied alone (24 h at 37°C) in pasteurized milk Martınez et al 2008 Lysis was inhibited when phage K was added to raw milk whey. This might be due to adsorption of whey proteins on S. aureus cells inhibited phage attachment Gill et al 2006 Adsorption of phage K was reduced in raw milk O'Flaherty et al 2005 Cronobacter sakazakii A cocktail composed of five phages prevented the growth of 35 of 40 test strains tested in 45 experimentally contaminated infant formula. Also, a dose of 10 8 PFU/mL eradicated the test strains from a liquid culture medium contaminated with both high and low concentrations (10 6 and 10 2 CFU mL −1 ) of the bacterial cells Zuber et al 2008 In experimentally contaminated infant formula phage concentration of 10 9 PFU/ml was the most effective and able to completely eradicate the target organism Kim et al 2007 and reduced levels of C. coli and C. jejuni in feces by 2 log CFU/g.…”

“…They should also display minimum transduction, a process wherein host DNA is packaged into phage heads, rather than phage DNA (Ikeda and Tomizawa 1965). Phages selected should be stable within the intended use environment (Gill and Abedon 2003). In addition to all these phages should also have a broad host range, however this limitation can be mitigated using phage cocktails (McIntyre et al 2007).…”

“…These phages are called temperate, and the cells that harbor a prophage are known as lysogenic. The lysogenic relationship between a temperate phage and its host bacterium provides a safe home to the temperate phage genome, blocks replication of non-virulent homologous phages, and has the potential to alter the phenotype of the host cell (Gill and Abedon 2003).…”

Characterization of a Novel Panton-Valentine Leukocidin (PVL)-Encoding Staphylococcal Phage and Its Naturally PVL-Lacking Variant

“…The presence of an integrase gene and toxin genes in phage genomes leads to poor antibacterial efficacy and safety concerns, respectively. Consequently, only virulent phages (or temperate phages replicating on indicator strains) lacking any virulence factor have been used as biocontrol agents against S. aureus in foods (13)(14)(15)(16) and in animal models (17)(18)(19). In an effort to isolate new virulent phages to control Staphylococcus contaminations in dairy products, we analyzed raw milk samples for the presence of staphylococcal phages.…”