The pathogenesis of persistence: the effects of delivery site and biofilm formation by Mycoplasma ovipneumoniae

Abstract

Mycoplasma ovipneumoniae is a respiratory pathogen in domestic sheep that is typically associated with polymicrobial pneumonia. M. ovipneumoniae infections can lead to decreased weight gains, increased susceptibility to secondary pathogens, and increased lamb mortality. Current understanding of how M. ovipneumoniae affects its host in the absence of a secondary pathogen remains limited, as are the site-specific effects of infection. M. ovipneumoniae infections have been observed to cause persistent infections that resist both the immune system and antimicrobial treatment. Related organisms in the Mycoplasmataceae family evade the immune system and treatment through the formation of a biofilm. Biofilms are formed of aggregated bacteria that adhere to a surface and form a protective extracellular polymeric matrix. However, it is difficult to assess the impacts of Mycoplasma biofilm formation on antibiotic susceptibility due to limitations in current methods for antibiotic susceptibility testing (AST). Additionally, whether M. ovipneumoniae can form a biofilm remains a matter of debate. This work aims to address these issues by investigating how M. ovipneumoniae infection alone can influence pathogenesis, and how infection with a secondary pathogen can vary due to the site of initial administration. It also seeks to address limitations in the current AST methods for Mycoplasma biofilms and to better characterize M. ovipneumoniae biofilm formation capacity. In four related publications, we first show that M. ovipneumoniae alone causes asymptomatic infections that cannot be cleared with antibiotics. We next compare how infection in the lower respiratory tract results in increased pathogenesis with a secondary pathogen compared to the upper respiratory tract. We then developed a method for AST of Mycoplasma spp. biofilms to compare the changes in resistance observed between planktonic cells and biofilms. Finally, we show robust M. ovipneumoniae biofilm formation in vitro, and associated increases in antibiotic tolerance and cellular survival. These observations suggest that infection with M. ovipneumoniae alone results in asymptomatic infection, while coinfection with M. haemolytica in the lower respiratory tract leads to more severe disease. We conclude that the observed antibiotic treatment failure in vivo was caused by M. ovipneumoniae biofilm formation that led to increased antibiotic tolerance.