Comparative Genomic Analysis of Acanthamoeba Endosymbionts Highlights the Role of Amoebae as a "Melting Pot" Shaping the Rickettsiales Evolution

Abstract

Amoebae have been considered as a genetic "melting pot" for its symbionts, facilitating genetic exchanges of the bacteria that co-inhabit the same host. To test the "melting pot" hypothesis, we analyzed six genomes of amoeba endosymbionts within Rickettsiales, four of which belong to Holosporaceae family and two to Candidatus Midichloriaceae. For the first time, we identified plasmids in obligate amoeba endosymbionts, which suggests conjugation as a potential mechanism for lateral gene transfers (LGTs) that underpin the "melting pot" hypothesis. We found strong evidence of recent LGTs between the Rickettsiales amoeba endosymbionts, suggesting that the LGTs are continuous and ongoing. In addition, comparative genomic and phylogenomic analyses revealed pervasive and recurrent LGTs between Rickettsiales and distantly related amoeba-associated bacteria throughout the Rickettsiales evolution. Many of these exchanged genes are important for amoeba-symbiont interactions, including genes in transport system, antibiotic resistance, stress response, and bacterial virulence, suggesting that LGTs have played important roles in the adaptation of endosymbionts to their intracellular habitats. Surprisingly, we found little evidence of LGTs between amoebae and their bacterial endosymbionts. Our study strongly supports the "melting pot" hypothesis and highlights the role of amoebae in shaping the Rickettsiales evolution.

Keywords: Rickettsiales; amoeba endosymbionts; lateral gene transfer.

Fig. 1.

—Predominance of amoeba-associated bacteria in the best hits of Rickettsiales genes. The histogram on the bottom right represents the taxonomic distribution of the non α-proteobacterial best hits (x axis) for genes in the six amoeba endosymbionts (highlighted in red) and the 13 Rickettsiales representatives (y axis). The top 10 ranked non-α-proteobacterial species with the largest number of best hits are shown. The histogram is enlarged and shown at the center.

Fig. 2.

—Gene transfer network illustrating LGTs between Rickettsiales and distantly related amoeba-associated bacteria. Each edge represents an LGT route between the two nodes. Edge color corresponds to the number of gene families transferred between the two nodes. The size of the node is proportional to the number of laterally transferred gene families involving the nodes. The six amoeba endosymbionts are highlighted in red.

Fig. 3.

—Maximum likelihood phylogeny for the ferritin-like protein showing LGT between Rickettsiales endosymbionts and four amoeba species. The tree was rooted using midpoint rooting. Bootstrap values for internal nodes are shown beside them. The four amoeba species are highlighted in red.