Analysis of virulence of the emerging opportunistic human pathogen Achromobacter in a zebrafish embryo model

Background: Achromobacter spp. are emerging opportunistic pathogens of concern in people with cystic fibrosis (CF) and immunocompromised individuals. These bacteria are known for their intrinsic antibiotic resistance and ability to evade host immune defences. A more comprehensive understanding of the direct interaction between Achromobacter spp. and the host innate immune system is necessary for the development of effective intervention strategies. In this study, we used zebrafish embryos to investigate the pathogenic potential of a panel of clinical Achromobacter spp. isolates and analysed their interaction with the host innate immune system. Results: Seven CF isolates previously shown to cause distinct levels of cytotoxicity in cell culture, and virulence and inflammation in Galleria mellonella larvae and mice, respectively, were analysed in a zebrafish embryo model. One strain caused significant host mortality (87.5%) in zebrafish embryos by 4 days post intravenous injection. Mortality was associated with a strong increase in bacterial load and increased pro-inflammatory cytokine gene expression. Three strains were moderately virulent and induced 8.4 to 12.5% host mortality, with surviving embryos having a constant bacterial load until 4 days post infection, indicating bacterial persistence. The remaining three isolates persisted, but caused less than 3% mortality, and were cleared by 27% of infected embryos, which was associated with minimal induction of pro-inflammatory gene expression at 1 day post infection. To elucidate the role of macrophages, macrophage-depleted embryos were compared with macrophage-proficient embryos after infection with the least virulent strain (0% mortality). The absence of macrophages led to a strong increase in bacterial load and 34% mortality within 4 days, demonstrating a significant role for macrophages in controlling bacterial growth and limiting infection severity. Real time analysis revealed, however, that some isolates were able to survive and replicate in macrophages, thereby contributing to either persistent or pro-inflammatory infection. These results demonstrated both host-detrimental and host-beneficial roles for macrophages during infection. Conclusion: Zebrafish embryos represent a valuable model for studying Achromobacter-host interactions during both persistent and acute infection, particularly to study the role of immune cells and obtain insights into host-pathogen interactions in real-time. The zebrafish infection model will be a helpful tool for the discovery antimicrobial strategies against Achromobacter spp.