BACTERIOPHAGE THERAPY: IS THIS THE WAY TO GO WITH MASTITIS?

Discipline: mastitis; 

Bacteriophage (“phage”) therapy has been exploited for the control of bacterial diseases in fauna, flora and humans. However, the advent of antibiotic therapy lead to a cessation of most phage research. Recently, the problem of antibiotic resistance has rendered many commonly used antibiotics ineffective, thereby renewing interest in phage therapy as an alternative source of control. This is particularly relevant in the case of bovine mastitis, caused by strains such as Staphylococcus aureus. Antibiotic resistance (primarily towards penicillin and methicillin) by staphylococcal strains causing mastitis is regularly reported.

Phage therapy can provide a stable, effective and affordable system of mastitis control with little to no deleterious effect on the surrounding environment or the affected animal itself. However, although several studies have addressed the issue of bio-control of bovine mastitis using phages, results are variable. While some phage-based products have been commercialized for the treatment of S. aureus-associated infections in humans, no products have yet been formulated specifically for the strains responsible for bovine mastitis. If the reliability of phage therapy can be resolved, then phages may become a primary form of control for bovine mastitis and other bacterial diseases.

The present studies investigated the presence of S. aureus and its phages in a dairy environment, as well as the controlling ability of phage isolates against antibiotic-resistant strains of mastitic S. aureus. The primary goals of the studies were to isolate promising phages as potential control agents for the disease, to conduct bioassays in the laboratory on the selected phages and to conduct phage sensitivity assays to assess phage activity against different chemical and environmental stresses that may limit their effectiveness as controlling agents in the field.  

Twenty-eight phages were isolated and screened against four strains of S. aureus. Only six phages showed potential for further testing, based on their wide host range, high titres and common growth requirements. Optimal growth conditions for the host S. aureus strain was 37°C for 12hours. This allowed for optimal phage replication. At an optimal determined titre these phages were able to reduce live bacterial cell counts by 64-95%. In addition, all six phages showed controlling activity towards another 18 S. aureus strains that were isolated from different milk-producing regions during a farm survey. These six phages were named Sabp-P1, Sabp-P2, Sabp-P3, Sabp-P4, Sabp-P5 and Sabp-P6.

Sensitivity bioassays towards simulated environmental and formulation stresses were subsequently conducted on the six identified phages. Phages Sabp-P1, Sabp-P2 and Sabp-P3 showed the most stable replication rates at increasing temperatures (45-70°C). The effect of temperature on storage of phages showed that 4ºC was the minimum temperature at which phages could be stored without a significant reduction in their controlling and replication abilities. Furthermore, all phages showed varying levels of sensitivity to chloroform exposure, with Sabp-P5 exhibiting the highest level of reduction in activity (74%) in comparison to the other phages. All six phages showed optimal controlling ability at pH 6.0-7.0 and reduced activity at any pH above or below pH 6.0-7.0. Exposure of phages to varying glycerol concentrations (5-100%) produced variable results. All six phages were most stable at a glycerol concentration of 10-15%.

The results showed that three of the six isolated phages, Sabp-P1, Sabp-P2 and Sabp-P3, performed optimally during the laboratory assays and have considerable potential for applications to treat mastitis-infected dairy cattle. These and further laboratory evaluations will soon be conducted by the research team in a research programme funded by Milk SA. 

References:

Basdew, I. H., 2012. Biological and molecular characterization of South African bacteriophages infective against Staphylococcus aureus subsp. aureus Rosenbach 1884, causal agent of bovine mastitis. PhD Thesis, UKZN, Pietermaritzburg.

Basdew, I. H. & Laing, M. D. 2014. Stress sensitivity assays of bacteriophages associated with Staphylococcus aureus, causal organism of bovine mastitis. Afr. J. Microbiol. Res.8, 200-210