Protein and DNA Biosynthesis Demonstrated in Host Cell-Free Phagosomes Containing Anaplasma phagocytophilum or Ehrlichia chaffeensis in Axenic Media

Rickettsiae belong to the Anaplasmataceae family, which includes mostly tick-transmitted pathogens causing human, canine, and ruminant diseases. Biochemical characterization of the pathogens remains a major challenge because of their obligate parasitism. We investigated the use of an axenic medium for growth of two important pathogens-Anaplasma phagocytophilum and Ehrlichia chaffeensis-in host cell-free phagosomes. We recently reported that the axenic medium promotes protein and DNA biosynthesis in host cell-free replicating form of E. chaffeensis, although the bacterial replication is limited. We now tested the hypothesis that growth on axenic medium can be improved if host cell-free rickettsia-containing phagosomes are used. Purification of phagosomes from A. phagocytophilum- and E. chaffeensis-infected host cells was accomplished by density gradient centrifugation combined with magnet-assisted cell sorting. Protein and DNA synthesis was observed for both organisms in cellfree phagosomes with glucose-6- phosphate and/or ATP. The levels of protein and DNA synthesis were the highest for a medium pH of 7. The data demonstrate bacterial DNA and protein synthesis for the first time in host cell- free phagosomes for two rickettsial pathogens. The host cell support-free axenic growth of obligate pathogenic rickettsiae will be critical in advancing research goals in many important tick-borne diseases impacting human and animal health.

Automated summary

The study explores the use of an axenic medium for the growth of two important rickettsiae pathogens in host cell-free phagosomes. It demonstrates the synthesis of bacterial DNA and protein in host cell-free phagosomes for the first time, providing new insights into the replication and growth of these pathogens. The findings highlight the importance of host cell support-free axenic growth in advancing research on tick-borne diseases affecting human and animal health.