Reduced and Nonreduced Genomes in Paraburkholderia Symbionts of Social Amoebas

Abstract

The social amoeba Dictyostelium discoideum is a predatory soil protist frequently used for studying host-pathogen interactions. A subset of D. discoideum strains isolated from soil persistently carry symbiotic Paraburkholderia, recently formally described as P. agricolaris, P. bonniea, and P. hayleyella. The three facultative symbiont species of D. discoideum present a unique opportunity to study a naturally occurring symbiosis in a laboratory model protist. There is a large difference in genome size between P. agricolaris (8.7 million base pairs [Mbp]) versus P. hayleyella and P. bonniea (4.1 Mbp). We took a comparative genomics approach and compared the three genomes of D. discoideum symbionts to 12 additional Paraburkholderia genomes to test for genome evolution patterns that frequently accompany host adaptation. Overall, P. agricolaris is difficult to distinguish from other Paraburkholderia based on its genome size and content, but the reduced genomes of P. bonniea and P. hayleyella display characteristics indicative of genome streamlining rather than deterioration during adaptation to their protist hosts. In addition, D. discoideum-symbiont genomes have increased secretion system and motility genes that may mediate interactions with their host. Specifically, adjacent BurBor-like type 3 and T6SS-5-like type 6 secretion system operons shared among all three D. discoideum-symbiont genomes may be important for host interaction. Horizontal transfer of these secretion system operons within the amoeba host environment may have contributed to the unique ability of these symbionts to establish and maintain a symbiotic relationship with D. discoideum. IMPORTANCE Protists are a diverse group of typically single cell eukaryotes. Bacteria and archaea that form long-term symbiotic relationships with protists may evolve in additional ways than those in relationships with multicellular eukaryotes such as plants, animals, or fungi. Social amoebas are a predatory soil protist sometimes found with symbiotic bacteria living inside their cells. They present a unique opportunity to explore a naturally occurring symbiosis in a protist frequently used for studying host-pathogen interactions. We show that one amoeba-symbiont species is similar to other related bacteria in genome size and content, while the two reduced-genome-symbiont species show characteristics of genome streamlining rather than deterioration during adaptation to their host. We also identify sets of genes present in all three amoeba-symbiont genomes that are potentially used for host-symbiont interactions. Because the amoeba symbionts are distantly related, the amoeba host environment may be where these genes were shared among symbionts.

Keywords: Burkholderia; Dictyostelium; genome reduction; protist; symbiosis.

FIG 1

Hive plot of whole-genome comparisons of D. discoideum-symbiont genomes. Locally colinear blocks between pairs of genomes are shown as bands that connect the axes (genomes). Only blocks above the median size are shown. Alignment of locally colinear blocks are distinguished between forward (blue) and reverse (purple) orientation. Axes are oriented center out, and boundaries between chromosomes are shown as ticks.

FIG 2

Comparison of reduced (red) and nonreduced (turquoise) genomes in terms of their functional compositions in nonmetric multidimensional space. The contributions of COG categories are projected with minor adjustments to avoid overlap with other features (pagri, P. agricolaris; pbonn, P. bonniea; phayl, P. hayleyella; pcale, P. caledonica; pfung, P. fungorum, pmega, P. megapolitana, pphem, P. phenazinium; pphex, P. phenoliruptrix; pphym, P. phymatum; pphyt, P. phytofirmans; psart, P. sartisoli; pspre, P. sprentiae; ptera, P. terricola; ptere, P. terrae; pxeno, P. xenovorans) (COG categories: J, Translation, ribosomal structure and biogenesis; A, RNA processing and modification; K, Transcription; L, Replication, recombination and repair; B, Chromatin structure and dynamics; D, Cell cycle control, cell division, chromosome partitioning; Y, Nuclear structure; V, Defense mechanisms; T, Signal transduction mechanisms; M, Cell wall/membrane/envelope biogenesis; N, Cell motility; Z, Cytoskeleton; W, Extracellular structures; U, Intracellular trafficking, secretion, and vesicular transport O, Posttranslational modification, protein turnover, chaperones; X, Mobilome: prophages, transposons; C, Energy production and conversion; G, Carbohydrate transport and metabolism; E, Amino acid transport and metabolism; F, Nucleotide transport and metabolism; H, Coenzyme transport and metabolism; I, Lipid transport and metabolism; P, Inorganic ion transport and metabolism; Q, Secondary metabolites biosynthesis, transport and catabolism; R, General function prediction only; S, Function unknown.

FIG 3

Representative individual COGs belonging to categories Cell motility (a), Transcription (b), Carbohydrate transport and metabolism (c), and Inorganic ion transport and metabolism (d) that were significantly overrepresented (a) or underrepresented (b–d) in the reduced genomes of D. discoideum symbionts. Contours of abundances are superimposed on the nonmetric multidimensional space from Fig. 2. P. bonniea and P. hayleyella are shown as red points to the left, while P. agricolaris is distinguished from the other genomes (white) as a turquoise point. Lighter blue contour lines indicate higher abundance compared to darker blue lines.

FIG 4

Core genes divided into the hypothesis that best predicts their patterns of molecular evolution. Core genes included genes evolving under stronger selective constraints with significantly lower dN/dS in genomes of symbionts of D. discoideum or other eukaryotes (“symbiotic” and “dicty”) and genes showing evidence of relaxed selective constraints with significantly higher dN/dS in the reduced genomes of P. bonniea and P. hayleyella (“reduced”). P. bonniea and P. hayleyella genes are included in the groups: symbiont, D. discoideum-symbiont, and reduced genome within each hypothesis.

FIG 5

The abundances of secretion systems detected in D. discoideum-symbiont genomes and other Paraburkholderia. For the type 4 secretion system, only protein secretion (as opposed to conjugation-related) T4SS abundances are shown. The phylogeny is a species tree of the 15 Paraburkholderia genomes we examined, reduced from the larger species tree in Brock et al. (10).

FIG 6

Type 3 secretion systems and flagella categorized using the conserved component genes sctJ (inner membrane ring; IPR003282), sctN (ATPase; IPR005714), and sctV (export apparatus; IPR006302). Branch lengths were ignored to improve readability of the ASTRAL tree topology. T3SS categories precede the name of the operon (e.g., “8 Pand7” is operon Pand7 belonging to category 8) downloaded from T3Enc database v1.0 (Hu et al. [135]). Tip labels for T3SS in the three D. discoideum-symbiont genomes, B. mallei (BMAA), or B. pseudomallei (BPSS) are shown in bold font face with gene IDs for ease of cross-reference. D. discoideum-symbiont genome T3SS operons are marked with a square symbol, and the clade containing the shared T3SS operon is shaded.

FIG 7

Type 6 secretion systems categorized using the conserved component genes tssB (sheath; COG3516), tssC (sheath; COG3517), and tssF (baseplate; COG3519). Branch lengths were ignored to improve readability of the ASTRAL tree topology. T6SS categories precede the name of the strain to which the operon belongs (e.g., “ii Francisella novicida U112” belongs to category ii), downloaded from SecReT6 database v3.0 (Li et al. [139]). Tip labels for T6SS in the three D. discoideum-symbiont genomes, B. mallei (BMAA), or B. pseudomallei (BPSS, BPSL) are shown in bold font face with gene IDs for ease of cross-reference. D. discoideum-symbiont genome T3SS operons are marked with a square symbol, and the clade containing the shared T6SS operon is shaded.