Multi-omics characterisation of the virulence and inflammatory potential of Bacillus thuringiensis in an in vitro...

The soil bacterium Bacillus thuringiensis (Bt) can produce spores and crystals composed of toxins with insecticidal activity. Due to these properties, some Bt strains have been developed as biopesticides for pest control, offering an alternative to chemical insecticides. However, Bt strains belong to the Bacillus cereus (Bc) group and share some virulence genes encoding diarrhoeal enterotoxins such as Hemolysin BL (Hbl), Non-haemolytic enterotoxin (Nhe) and Cytotoxin K (Cyt-K2). These toxins are known to be involved in foodborne outbreaks (FBOs) caused by Bc. For this reason, it has been suggested that Bt may also cause FBOs, making it crucial to evaluate their virulence potential and effects on human health. To address this, differentiated human intestinal Caco-2 cells, which mimic the intestinal barrier, were treated with serial dilutions of culture supernatants from five Bt strains used as biopesticides. No cytotoxicity was observed with any of the Bt supernatants after 24 h treatment in contrast to Bacillus cytotoxicus (Bcyt) supernatants. However, interleukin 8 (IL-8) secretion increased sharply for Bt as well as Bcyt, suggesting a potential for chronic inflammation. To investigate the molecular effects of Bt supernatants on Caco-2 cells, untargeted proteomics and metabolomics were performed on cell media and lysates using two non-cytotoxic concentrations. Both approaches highlighted common perturbations on oxidative phosphorylation and lipid metabolism for exposure to Bt or Bcyt supernatants. Proteins linked to acute inflammation decreased only with Bcyt exposure. Bt supernatants, but not Bcyt supernatants caused shedding of CD59 and CD55, two membrane-bound complement inhibitors, into the culture medium. This demonstrated that Bt supernatants could increase the vulnerability of intestinal cells to their own immune system. Proteomics also revealed the heterogeneous toxin abundances in the supernatants of the different Bt subspecies, highlighting the need to assess each strain individually when assessing the risks associated with their use as biopesticides.