High level of intergenera gene exchange shapes the evolution of haloarchaea in an isolated Antarctic lake Matthew Z. DeMaerea, Timothy J. Williamsa, Michelle A. Allena, Mark V. Browna,b, John A. E. Gibsonc, John Richa, Federico M. Lauroa, Michael Dyall-Smithd, Karen W. Davenporte, Tanja Woykef, Nikos C. Kyrpidesf, Susannah G. Tringef, and Ricardo Cavicchiolia,1 aSchool of Biotechnology and Biomolecular Sciences, The University of New South Wales, Sydney, NSW 2052, Australia; bEvolution and Ecology Research Centre, The University of New South Wales, NSW 2052, Australia; cInstitute of Marine and Antarctic Studies, University of Tasmania, Hobart, TAS 7001, Australia; dCharles Sturt University, Wagga Wagga, NSW 2678, Australia; eDepartment of Energy Joint Genome Institute Bioscience Division, Los Alamos National Laboratory, Los Alamos, NM 87545; and fDepartment of Energy Joint Genome Institute, Walnut Creek, CA 94598 Edited by W. Ford Doolittle, Dalhousie University, Halifax, NS, Canada, and approved September 5, 2013 (received for review April 17, 2013) Deep Lake in Antarctica is a globally isolated, hypersaline system have noted the ability of H. lacusprofundi, and a recent isolate, that remains liquid at temperatures down to −20 °C. By analyzing tADL (14), to form aggregates and biofilms at either high or low metagenome data and genomes of four isolates we assessed ge- temperatures (13, 15, 16), the only studies addressing adaptive nome variation and patterns of gene exchange to learn how the responses are those linking the production of unsaturated diether lake community evolved. The lake is completely and uniformly membrane lipids to cold adaptation in H. lacusprofundi (3, 17). dominated by haloarchaea, comprising a hierarchically structured, Various mechanisms that alter genetic composition have been low-complexity community that differs greatly to temperate and reported in haloarchaea, including archaeal viruses, conjugative plasmids, genome rearrangements mediated by transposons, and tropical hypersaline environments. The four Deep Lake isolates fi represent distinct genera (∼85% 16S rRNA gene similarity and signi cant levels of gene exchange via the formation of hetero- ∼ diploids followed by homologous recombination (18–25). All 73% genome average nucleotide identity) with genomic charac- fi teristics indicative of niche adaptation, and collectively account for these studies have been con ned to temperate or tropical sys- tems, not polar environments. ∼72% of the cellular community. Network analysis revealed a re- For some bacteria, the relationship between recombination EVOLUTION markable level of intergenera gene exchange, including the shar- frequency and sequence divergence appears to be log-linear (26, ing of long contiguous regions (up to 35 kb) of high identity ∼ Halobacterium 27) but may be as much as two orders of magnitude higher in ( 100%). Although the genomes of closely related , haloarchaea, such as for Haloferax volcanii and Haloferax medi- Haloquadratum Haloarcula > , and ( 90% average nucleotide iden- terranei, which readily undergo cell fusion and DNA exchange tity) shared regions of high identity between species or strains, the (25, 28). Forces driving speciation in microbes include niche four Deep Lake isolates were the only distantly related haloarch- adaptation, selective sweeps, genetic drift, recombination, and aea to share long high-identity regions. Moreover, the Deep Lake geographic isolation (29). However, it is unclear how these forces high-identity regions did not match to any other hypersaline en- would maintain species homogeneity or bring about lineages vironment metagenome data. The most abundant species, tADL, when gene flow is high, as is the case in haloarchaea. Although appears to play a central role in the exchange of insertion sequen- ces, but not the exchange of high-identity regions. The genomic Significance characteristics of the four haloarchaea are consistent with a lake ecosystem that sustains a high level of intergenera gene exchange Horizontal gene exchange across species boundaries is con- while selecting for ecotypes that maintain sympatric speciation. The sidered infrequent relative to vertical inheritance that main- peculiarities of this polar system restrict which species can grow and tains species coherence. However, haloarchaea living in provide a tempo and mode for accentuating gene exchange. hypersaline environments take a more relaxed approach to gene exchange. Here we demonstrate that in Deep Lake, Antarctica, mobile genetic elements | Antarctic haloarchaea | saltern | haloarchaea exchange DNA between distinct genera, not just fragment recruitment | BJ1 virus species, with some of the DNA being long (up to 35 kb) and virtually 100% conserved. With extremely low cell division rates eep Lake (DL) is an extremely cold and hypersaline envi- in the cold (e.g., six generations per year), the remarkable extent Dronment that has the distinction of being the least pro- of lateral exchange could conceivably homogenize the pop- ductive lake ever recorded (1–3). The lake is a marine-derived ulation. It is therefore equally notable that despite the demon- system in the Vestfold Hills, East Antarctica, having been iso- strated capacity for exchange, different genera are maintained, lated from the Southern Ocean by isostatic rebound of the their coexistence being linked to genomic differences conferring continent ∼3,500 y BP (2, 4, 5) (SI Appendix, Fig. S1). The ecotype distinctions that enable niche adaptation. temperature exceeds 0 °C only in the top few meters for a few summer months per year, and it remains ice-free even in winter Author contributions: J.A.E.G. and R.C. designed research; T.J.W., M.V.B., J.R., F.M.L., when temperatures drop to −40 °C (air) and −20 °C (throughout M.D.-S., T.W., N.C.K., S.G.T., and R.C. performed research; M.Z.D., T.J.W., M.A.A., M.V.B., the lake) (1, 6). Microbial diversity is extremely low, and is dom- K.W.D., and R.C. analyzed data; and M.Z.D. and R.C. wrote the paper. inated by members of the haloarchaea (7). Within this ecosystem, The authors declare no conflict of interest. the putative primary producer is the green alga Dunaliella sp., This article is a PNAS Direct Submission. which is found at low biomass concentrations (8–10). Data deposition: The genome data reported in this paper have been deposited in the From DL, Halorubrum lacusprofundi, the first member of the National Center for Biotechnology Information BioProject database, www.ncbi.nlm.nih. Archaea domain isolated from a cold environment (11), has been gov/bioproject [accession nos. PRJNA53493 (tADL), PRJNA58807 (HI), PRJNA72619 (DL31), H. lacusprofundi and PRJNA75121 (DL1)]; and the metagenome data have been deposited in the Genomes formally described (12). In the laboratory, OnLine database, www.genomesonline.org [accession nos. Gs0000582 (13m_0.1), grows across a wide range of temperatures from −1°Cto42°C, Gs0000585 (24m_0.8), Gs0000586 (SSU rRNA pyrotag data), Gs0000587 (24m_0.1), with fastest growth rate occurring at ∼33 °C (12, 13). It is capable Gs0000592 (36m_pooled), Gs0000594 (24m_3.0), and Gs0000595 (5mRS_0.1)]. of using a number of different carbon sources including glucose, 1To whom correspondence should be addressed. E-mail: [email protected]. mannose, acetate and ethanol, providing it with a seemingly This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10. versatile heterotrophic metabolism (12). Aside from studies that 1073/pnas.1307090110/-/DCSupplemental. www.pnas.org/cgi/doi/10.1073/pnas.1307090110 PNAS | October 15, 2013 | vol. 110 | no. 42 | 16939–16944 Downloaded by guest on September 27, 2021 selection is important for speciation, it has been argued that the fraction, compared with the relatively even distribution of tADL rate of genomic recombination may be a key determinant when across all size fractions (SI Appendix,Fig.S6). considering sympatric speciation. Modeling has shown that high From the analysis of eight million metagenomic 454 reads recombination rates are necessary to generate a new species against all available complete and draft genomes, the four DL when an ecological trait inferring increased fitness is controlled genomes recruited the largest number of reads (94.4% of all by many adaptive loci (30). However, these same high levels of recruited reads) and 28.3% of all reads. Similarly, as a pro- recombination may also increase the number of intermediate portion of Illumina reads, the four DL genomes represented (suboptimal) genotypes and reduce “completeness” of specia- 50% of all reads. The lower rate of recruitment for 454 may be tion. The dynamics of this sympatric evolution hypothesis have explained by the technology’s longer read length and therefore not been examined in real populations. increased rejection of partial degenerate sequence alignment set The DL system is geographically very isolated, its organismal by the stringent minimum alignment coverage (98%). The un- growth rates are very low, and the combination of cold plus hy- assigned metagenome data could derive from viruses and/or persalinity may be expected to promote the physical process of eucarya that are not in the National Center for Biotechnology DNA transformation to levels that exceed those of warm hyper- Information database (SI Appendix). saline environments. In this context, we examined the genomes of four recent haloarchaeal isolates and metagenome data from Characteristics of the Nine Replicons of the Four Genomes. Second- DL to assess genome variation and patterns of gene exchange ary replicons in DL31, H. lacusprofundi and DL1 had consis- and derive an understanding of how this unique Antarctic lake tently lower codon adaptation (CAI) and codon bias (CBI) community evolves. indexes (34, 35) than the primary replicons. Additionally, the primary replicons of tADL and DL1 possessed extended regions Results with low CAI/CBI. The low CAI/CBI regions often coincided fi with strong variations in FR and increased density of mobile DL Microbial Community Composition. From biomass lter frac- A B SI Appendix tionated (20–3.0, 3.0–0.8, 0.8–0.1 μm) from four depths of DL (5, elements (Fig.