Streptococcus iniae biofilm formation enhances environmental persistence and resistance to antimicrobials and disinfectants

The globally distributed bacterium Streptococcus iniae is responsible for outbreaks of disease resulting in high mortality in a wide range of economically important freshwater and marine fish species. Despite the significance of S. iniae, our understanding of its transmission and infection dynamics remains incomplete. Biofilms are important for the survival and pathogenesis of many bacteria, but there is a paucity of information on their role in the ex-host persistence of S. iniae. This study aimed to compare biofilm formation by isolates representing different S. iniae genotypes and to investigate the effect of biofilm formation on environmental persistence and resistance to common disinfectants and antimicrobials. Eleven clinical isolates of S. iniae representing 4 distinct genetic groups and diverse host types were assessed for their ability to form biofilms. Planktonic bacteria or mature biofilms were exposed to in vitro aquatic microcosms of different temperatures to quantify the number of culturable bacteria in each system over time. The minimum biofilm eradication concentration (MBEC) assay® system was used to determine biofilm resistance to 18 antimicrobials and 4 disinfectants commonly used in food producing animals and aquaculture, respectively. All isolates formed biofilms within 72 h. Bacteria remained culturable notably longer in the biofilm form compared to the planktonic, with a significant impact from temperature and salinity (p < 0.05). The MBEC was higher than the planktonic minimal inhibitory concentration (MIC) for at least one isolate in 15 out of the 18 antimicrobials tested. The MBEC was also higher than the minimum biocidal concentration (MBC) for 11 out of 18 tested, including oxytetracycline and florfenicol, the two most common antimicrobials used against S. iniae infections in fish. While both forms were susceptible to disinfection by bleach, hydrogen peroxide and Virkon® Aquatic, treatment with povidone‑iodine did not eliminate biofilms. The ability of S. iniae to form resilient biofilms provides an effective mechanism for their persistence in the environment, which must be considered and further researched to control this widespread pathogen.