Biofilms are a common cause of food contamination with undesirable bacteria, such as pathogenic bacteria. Staphylococcus aureus is one of the major bacteria causing food-borne diseases in humans. A study designed to determine the presence of S. aureus on food contact surfaces in dairy, meat, and seafood environments and to identify coexisting microbiota has therefore been carried out. A total of 442 samples were collected, and the presence of S. aureus was confirmed in 6.1% of samples. Sixty-three S. aureus isolates were recovered and typed by random amplification of polymorphic DNA (RAPD). Profiles were clustered into four groups which were related to specific food environments. All isolates harbored some potential virulence factors such as enterotoxin production genes, biofilm formation-associated genes, antibiotic resistance, or lysogeny. PCR-denaturing gradient gel electrophoresis (PCR-DGGE) fingerprints of bacterial communities coexisting with S. aureus revealed the presence of bacteria either involved in food spoilage or of concern for food safety in all food environments. Food industry surfaces could thus be a reservoir for S. aureus forming complex communities with undesirable bacteria in multispecies biofilms. Uneven microbiological conditions were found in each food sector, which indicates the need to improve hygienic conditions in food processing facilities, particularly the removal of bacterial biofilms, to enhance the safety of food products.

Staphylococcal biofilms are a major concern in both clinical and food settings because they are an important source of contamination. The efficacy of established cleaning procedures is often hindered due to the ability of some antimicrobial compounds to induce biofilm formation, and to the presence of persister cells, a small bacterial subpopulation that exhibits multidrug tolerance. Phage lytic enzymes have demonstrated antimicrobial activity against planktonic and sessile bacteria. However, their ability to lyse and/or select persister cells remains largely unexplored so far. In this work, the lytic activity of the endolysin LysH5 against Staphylococcus aureus and Staphylococcus epidermidis biofilms was confirmed. LysH5 reduced staphylococcal sessile cell counts by 1–3 log units, compared with the untreated control, and sub-inhibitory concentrations of this protein did not induce biofilm formation. LysH5-surviving cells were not resistant to the lytic activity of this protein, suggesting that no persister cells were selected. Moreover, to prove the lytic ability of LysH5 against this subpopulation, both S. aureus exponential cultures and persister cells obtained after treatment with rifampicin and ciprofloxacin were subsequently treated with LysH5. The results demonstrated that besides the notable activity of endolysin LysH5 against staphylococcal biofilms, persister cells were also inhibited, which raises new opportunities as an adjuvant for some antibiotics.

Phage therapy is a promising option for fighting against staphylococcal infections. Two lytic phages, vB_SauM_phiIPLA-RODI (phiIPLA-RODI) and vB_SepM_phiIPLA-C1C (phiIPLA-C1C), belonging to the Myoviridae family and exhibiting wide host ranges, were characterized in this study. The complete genome sequences comprised 142,348 bp and 140,961 bp and contained 213 and 203 open reading frames, respectively. The gene organization was typical of Spounavirinae members, with long direct terminal repeats (LTRs), genes grouped into modules not clearly separated from each other, and several group I introns. In addition, four genes encoding tRNAs were identified in phiIPLA-RODI. Comparative DNA sequence analysis showed high similarities with two phages, GH15 and 676Z, belonging to the Twort-like virus genus (nucleotide identities of >84%); for phiIPLA-C1C, a high similarity with phage phiIBB-SEP1 was observed (identity of 80%). Challenge assays of phages phiIPLA-RODI and phiIPLA-C1C against planktonic staphylococcal cells confirmed their lytic ability, as they were able to remove 5 log units in 8 h. Exposure of biofilms to phages phiIPLA-RODI and phiIPLA-C1C reduced the amount of adhered bacteria to about 2 log units in both monospecies and dual-species biofilms, but phiIPLA-RODI turned out to be as effective as the mixture of both phages. Moreover, the frequencies of bacteriophage-insensitive mutants (BIMs) of Staphylococcus aureus and S. epidermidis with resistance to phiIPLA-RODI and phiIPLA-C1C were low, at 4.05 × 10 ⁻⁷ ± 2.34 × 10 ⁻⁹ and 1.1 × 10 ⁻⁷ ± 2.08 × 10 ⁻⁹ , respectively. Overall, a generally reduced fitness in the absence of phages was observed for BIMs, which showed a restored phage-sensitive phenotype in a few generations. These results confirm that lytic bacteriophages can be efficient biofilm-disrupting agents, supporting their potential as antimicrobials against staphylococcal infections.