Environmental and clinical isolates of Herbaspirillum induce pulmonary infection in mice and its secretome is cytotoxic to human lung cells

Introduction. In recent years, the Herbaspirillum genus has emerged as a pathogen in healthcare-related infections and has became stablished as an opportunistic pathogen. Hypothesis/Gap Statement. Little is known about the pathogenesis induced by Herbaspirillum genus. Aim. To evaluate the cytotoxic effects of genus Herbaspirillum, its ability to adhere to lung human cells and the ability of environmental and clinical strains of Herbaspirillum to induce pneumonia in mice. Methodology. Environmental and clinical isolates of Herbaspirillum were examined for their cytotoxic effects on the Calu-3 cell lineage. Cytotoxic activity of secretome was tested using MTT/neutral red assays and cell morphology analysis. Herbaspirillum adhesion on Calu-3 cells was assessed using bright-field microscopy and cell-associated bacteria were counted. A mouse model of acute lung infection was done using a clinical and an environmental strain. Adult male mice were used, and the pneumonia was inducted by intra-tracheal inoculation of 10(9) or 10(9) bacteria. Mice weight variations were evaluated at the end of the experiment. Bronchoalveolar lavage was collected and evaluated for total and differential cytology. A histological examination of lungs was performed giving a histological score. Results. The secretomes of all the strains induced morphological alterations in cells, but only H. seropedicae SmR1 were cytotoxic in MTT and neutral red assays. Clinical strains of H. frisingense AU14459 and H. hutttiense subsp. huttiense AU11883 exhibited low adherence to lung cells, while SmR1 was non-adhesive. Following intratracheal inoculation, mice treated with 10(9) c.f.u. of the SmR1 and AU11883 strains lost 18 and 6% of their weight over 7 days, respectively, and presented moderate clinical signs. Infected mice showed inflammatory cell infiltration in the perivascular and peribroncheal/peribronchiolar spaces. Bronchoalveolar fluid of mice inoculated with SmR1 10(9) c.f.u. presented an increase in total leucocyte cells and in neutrophils population. Conclusion. These in vivo and in vitro results provide insights into how some Herbaspirillum strains cause infection in humans, providing a basis for the characterization of pathogenesis studies on this emerging infectious agent.