Invited presentation at the 7th Annual Argonne Soil Metagenomics Meeting, 22 October 2015. Not very polished.

1. Soil, lateral gene transfer,
and hybrid genomes
Robert Beiko
20 October 2015

2. A. microbe
Lateral gene transfer
http://genome.cbs.dtu.dk/staff/dave/MScourse/Lekt_11Feb2003c.html

3. In the gut
Butyrate synthesis in Lachnospiraceae and other
organisms
Meehan and Beiko (2014) Genome Biol Evol
Lachnospiraceae

4. LGT across habitats
Smillie et al. (2011) Nature
Within-site transfer rates are
highest in host-associated (i.e.,
human) habitats
AR genes are frequently
transferred BETWEEN habitats

5. Villegas-Torres et al. (2011) International Biodeterioration & Biodegradation
Forsberg et al. (2012) Science
Sorangium cellulosum So157-2
14.8 Mbp; 11,599 coding sequences
>1200 putative LGT acquisitions
Han et al. (2013) Sci Rep

6. An ecological view of genomes
Genes as individuals, Genomes as communities
Key concept mappings:
• Diversity: counts of genes / distribution across functional
categories
• Community: set of genes and their interactions
• Migration: lateral gene transfer

7. Metacommunity
Leibold et al., Ecol Lett, 2004
A set of local communities that are linked by dispersal
Species B
Species A
Habitat
Habitat
Habitat
Pattern and intensity
of migration for species A

8. Genome Metacommunity
Hypothesis
• Since…
• Genes are agents whose trajectories are not bound to
their host organisms
• Genes can evolve and take on new functional roles in
concert with other genes
• A genome can be viewed as a community of genes
• Related sets of genomes comprise a
metacommunity of genes

9. Genome Metacommunities
Boon et al., Fems Microbiol Rev, 2014
A set of genomes that are linked by LGT
Gene B
Gene A
Genome
Genome
Genome
Pattern and intensity
of LGT for gene A

10. Genome Metacommunities
Boon et al., Fems Microbiol Rev, 2014
Related to the pan-genome, but not
restricted to specific taxonomic groups
Why is a given gene present in a given
genome at a given time?
How are functional roles partitioned
across a community?

11. Soil thinking
How important is LGT in soil communities?
Does it make sense to think of gene metacommunities
in the soil context?
Lots of LGT
YES
Minimal LGT
NO

12. The procedure
• In the absence of a coherent set of known genomes
from a given habitat…
1. Identify an interesting sample
2. Select genomes with very high marker-gene (i.e.,
16S) similarity to sequences in the sample
(gOTUs)
3. Mine genomes for evidence of LGT, examine
patterns of connectivity

13. Conclusions
• Positive relationship between
abundance, diversity and pH
• Specific relationships between different
bacterial (notably Acidobacteria) and
fungal groups vs. pH
• Fungal OTUs appear to tolerate wider pH
ranges
(1)
Chosen sample:
http://metagenomics.anl.gov/?page=MetagenomeOverview&metagenome=4455674.3#org_ref (pH = 4.1)

14. Meet the Sample (MG-RAST)
1277 rRNA
gene sequences

15. Meet the Sample (Matching genomes)
99% 16S identity (e-value < 1e-20):
1211 – No match
Bradyrhizobiaceae: 61
Pseudomonas: 2
Nocardioides: 1
Acidithiobacillus: 1
Cyanobium: 1
Total: 18 genomes covering 8 genera
97% 16S identity:
1100 – No match
Bradyrhizobiaceae: 77
Pseudoxanthomonas / Cycloclasticus: 25
Acidobacteria: 20
Other Proteobacteria: 48
Other: 10
Total: 114 genomes covering 74 genera
1277 rRNA gene sequences
(1)

16. gOTUs
16S sequence from sample
Rhodopseudomonas
palustris TIE 1
Rhodopseudomonas
palustris DX 1
Rhodopseudomonas
palustris CGA009
Bradyrhizobium
japonicum USDA 6
99% 16S identity
97% 16S identity
141824_31298
Rhodopseudomonas
palustris HaA2
Rhodopseudomonas
palustris BisA53
Bradyrhizobium BTAi1Nitrobacter
winogradskyi
Oligotropha
carboxidovorans
Agromonas
oligotrophica
Bradyrhizobium
ORS278

17. Weird gOTUs
141824_229613
Bordetella pertussis
Bordetella
bronchiseptica
Bordetella
parapertussis
Gross et al., 2008

18. Homology search
• Compare proxy genomes against nr database
• Identify interesting patterns:
• Unusual best matches (e.g., best nonself match is to a
completely different group)
• Patchy distributions, phylogenetic trees
• Linked sets of genes: co-transfer?
• Implicated biological processes?

19. https://microbewiki.kenyon.edu/index.php/Acidithiobacillus_ferrooxidans

20. Beiko (2011) Biol Direct
504 gene trees in which A. ferrooxidans has a unique genus as partner
Not shown: 795 genes w/multiple partners
Also not shown: 333 other trees with less frequent, unique partner genera
Split by 16S; reunited by genome sequencing?

21. Genome 1 – Acidithiobacillus ferrivorans
(renaming of A. ferrooxidans)
3093 predicted proteins / 3035 with homology
matches
Observed / Predicted capabilities:
• Facultatively anaerobic
• Psychrotolerant
• Optimal pH = 2.5
• Oxidation of iron and inorganic sulfur
• Carbon fixation, nitrate reduction
• Trehalose synthesis
• “Bioleaching”
Liljeqvist et al. (2011) J Bacteriol

22. Genome 1 – Acidithiobacillus ferrivorans
Best nonself match is to…
(273 non-Acidithiobacillus)

23. Mobile element signatures
dominate
• 14 x restriction system-associated
• 8 x transposase
• 8 x transcriptional regulators (incl CopG, TetR)
• Other resistance (LacZ, bleomycin, …)
• Integrase, reverse transcriptase, toxin/antitoxin,
bacteriocin, …
• Nitrate reductase & related
• >90 unknown

24. 1877 found in other
Acidithiobacillus + other
genera
Best non-Acidithiobacillus
match is to…
(only 11 Acidobacteria!)

25. https://www.jasondavies.com/wordcloud/#

26. Acidobacterial connections
• short-chain dehydrogenase/reductase SDR
• HNH endonuclease
• Glycoside hydrolase family 8 (x3)
• RES domain protein
• Transposase x 5

27. Phylogenetic profiles
# of similar genes (evalue < 10-50)
Min 30 connections
Proteobacteria
Actinobacteria
Cyanobacteria
Planctomycetes
Acidobacteria
Bacteroidetes
Acidithiobacillus
Key observations
• Connections to many other groups,
mostly Proteobacteria (not surprising)
• No between-group connections outside
Proteobacteria at this threshold
• Acidithiobacillus as hub rather than part
of gene-exchange community?
65

28. Mutual information-based network
(do groups co-occur > random?)
Acidithiobacillus
Gammaproteobacteria
Alpha/Betaproteobacteria
Key observations
• Connections mostly predictable by
phylogeny
• Again, no interesting partners outside of
Proteobacteria
• However, many connections between
Alpha/Betaproteobacteria

29. Phosphate ABC
transporters
gi 343775109
periplasmic
(eval < 10-100)
gi 343775110
inner membrane
subunit PstC
(eval < 10-50)
gi 343775111
inner membrane
subunit PstA
(eval < 10-50)
Distribution:
• Acidithiobacillus
• Acidobacterium
• Alpha/Beta/Gamma
• Actinobacteria
• Firmicutes
Recurrent grouping of
Acidithiobacillus (Gamma)
Acidobacterium
Thiobacillus (Beta)
Defluviimonas (Alpha)
Salinisphaera (Gamma)

30. Genome 2 – Terriglobus roseus
Eichorst et al (2007) IJSEM
• “Group 1” acidobacterium
• Preferred pH: ~6
• Aerobic
• Catalase, carotenoids for defense against reactive oxygen
• Oligotrophic; can grow on a wide range of carbon sources
• 4245 protein-coding genes (2735 with nr matches, 558
species-specific)
Rousk et al. Eichorst et al.

31. Best nonself matches (183 non-Acidobacteria)
Multidrug resistance / cation efflux / prophage

32. Best matches outside Terriglobus

33. Phylogenetic profiles
Profiles are wider and more diverse for
Terriglobus than for Acidithiobacillus

34. LPS O-antigen
biosynthesis
gi 390412425
CDP-glucose
4,6-dehydratase
gi 390412426
glucose-1-phosphate
cytidylyltransferase
gi 390412427
“LPS biosynthesis
protein”
Distribution:
• Acidithiobacillus
• Acidobacteria
• Other proteobacteria
• Other
Flavobacteria
Spirochaetes
Spirochaetes
Cyanobacteria

35. Contrasting Acidithiobacillus vs
Terriglobus relationships:
Same partners, different dance
Compare profiles vs Streptomycetaceae (five strains
found in sample gOTU)
Acidithiobacillus Common Terriglobus
Polyphosphate
kinase
glucose-6-
phosphate 1-
dehydrogenase
Carbon monoxide
dehydrogenase
More glycolytic
enzymes
Heavy-metal
resistance / export
Multidrug
resistance
Ammonium
transporter
Catalase /
peroxidase
Exopolysaccaride

36. Conclusions
• Different layers of LGT:
• Very recent: mostly mobile elements (proxies unsuitable)
• Less recent (outside species / genus) (proxies potentially
more justifiable)
• Taxonomy is a pain
• What’s the story with gene metacommunities?
• Lots of LGT!
• Recurrent patterns of sharing among groups not evident
• Metacommunities at the pan-genome level?
• Need many isolate genomes from single samples

37. Technical impacts of LGT and
gene metacommunities
Metagenomic read assignment
• Recently acquired genes will still look like they belong
in the donor
• These are some of the most interesting genes!!
Functional prediction (e.g., PICRUSt)
• Phylogeny will fail to accurately predict the distribution
of these genes. Be very careful with extreme or poorly
characterized samples!
Phylogenetic beta diversity may be misleading

38. Key questions in LGT and gene
metacommunities
• Are gene-sharing networks:
• Random?
• Driven by shared location / habitat?
• Constrained by phylogenetic relatedness?
• Are shared genes:
• Neutral or adaptive?
• Driven by specific types of mobile element?