The problem: Most probiotics fail to persist in host tissues. Benefits are transient; reapplication is costly and often impractical.
Our solution: We select probiotics based on genomic signatures of host dependency: insertion sequence proliferation, pseudogenization of core genes, and genome restructuring. These hallmarks identify lineages undergoing an irreversible evolutionary transition toward host association. Such bacteria are intrinsically predisposed to long-term residency.
Proof of concept - Enhancing coral reef resilience with Ruegeria MC10: From more than 1,200 coral-associated bacterial isolates, we identified Ruegeria population MC10, a marine Roseobacter exhibiting these genomic signatures. In an eight-month field trial spanning a natural bleaching event, corals that received a nursery application of MC10 maintained significantly higher coloration and photosynthetic efficiency than placebo (autoclaved, filtered seawater) controls. Controlled experiments confirmed that this enhanced thermal resilience required live, metabolically active cells and could not be explained by transient mechanisms such as nutritional supplementation or priming effects.
Evolutionary trade-offs: Host dependency came at a cost: genomic decay (insertion sequence expansion, pseudogene proliferation) and reduced free-living growth. Yet this evolutionary trade-off also brought functional innovations. MC10 uniquely encodes siderophore clusters for iron scavenging, diverse glycosyltransferases, and exopolysaccharide biosynthesis pathways. It forms robust biofilms and, upon host exposure, undergoes a coordinated proteomic switch from motility to sessility. These traits enable stable host integration. The outcome: enhanced host benefit.
CORDAP support: This work is funded by CORDAP (Coral Research and Development Accelerator Platform), a G20-backed international initiative to fast-track research and development solutions to save the world's corals. Our project, “Next-generation probiotics for corals”, runs from March 2026 to February 2029.
Building a global platform: We are constructing a global catalog of localized probiotics for ecologically safe restoration, engineering delivery systems for nursery and in situ applications, and decoding probiotic-host interactions using advanced imaging, single-cell omics, and genome editing tools.
Broader applications: We are adapting the same evolutionary framework for shellfish aquaculture, where current probiotics fail during hatchery-to-ocean transition. Our evolution-guided solutions, selected for persistence, address this critical bottleneck for sustainable aquaculture.
Key published / submitted papers in this area:
M. Xie, N. Xiang, T. Liao, C.T. Cheung, Z. Xian, P. Li, C.H. Lee, W.Y. Tse, K.K. Tsang, C. Xu, K.E. Ho, Q. He, M. Dörr, H. Manns, X.Wang, D. Luo, R. Hayden, E. Chei, Z. Wan, P. Thompson, J. Brennan, R.S. Peixoto, G. Cui, S.E. McIlroy, A.P.Y. Chui, C.R. Voolstra, and H. Luo. 2026. Evolutionary genomics predicts probiotic persistence in corals. (under review, bioRxiv)
M. Xie, C. Xu, N. Xiang, T. Liao, X. Liu, Z. Liu, X. Feng, Q. He, Z. Liang, W. Wang, Y. Dai, L.Yan, C. Pogoreutz, L. Barra, S.W.N. Au, L. Jiang, C.R. Voolstra, and H. Luo. 2026. From genomic decay to functional advantage: Trait-based evidence for long-term host residency of a next-generation coral probiotic. (under review, bioRxiv)
N. Xiang, T. Liao, M. Xie, Z. Wang, C.H. Mak, X. Tang, S.E. McIlroy, B. Thibodeau, C.R. Voolstra, and H. Luo. 2026. Decoding coral resistance to eutrophication: association of hyper-efficient denitrifiers as key microbial allies. Nature Communications (in press)
For decades, scientists assumed that Prochlorococcus, the tiny cyanobacterium responsible for producing 10% of the oxygen we breathe, evolved under the relentless force of natural selection. Because these cells are so abundant in the ocean, the logic went, their populations must be enormous, making genetic drift (the random fluctuation of gene frequencies) essentially irrelevant. This assumption gave rise to the “genome streamlining” theory, which proposes that selection actively purges unnecessary DNA to create efficient, minimal genomes. Our laboratory has spent the past several years questioning this narrative, and in doing so, we have uncovered a far more complex and fascinating story that weaves together Earth’s geological history with the fundamental principles of evolutionary genetics.
The first clue came from an unexpected place: the types of mutations that accumulated as Prochlorococcus genomes shrank. By distinguishing between conservative amino acid changes (those that preserve protein chemistry) and radical changes (those that alter it), we discovered that early genome reduction was accompanied by an excess of these more disruptive mutations. This pattern is the fingerprint of population collapse, not efficient selection. But what could have caused such a collapse in the ocean's most successful photosynthetic organism? The answer, we discovered, lies in one of the most dramatic episodes in Earth’s history: the Neoproterozoic Snowball Earth events, when global glaciation wrapped the planet in ice for millions of years. During this time, Prochlorococcus populations were squeezed into isolated refugia, such as brine channels within sea ice and cryoconite holes on glacier surfaces, where they experienced severe bottlenecks that left permanent marks in their genomes.
If ancient Prochlorococcus populations were indeed this small, we reasoned, then modern populations might carry echoes of this history. Through a painstaking three-year mutation accumulation experiment, tracking 141 lines of Prochlorococcus through more than a thousand generations, we directly measured their spontaneous mutation rate for the first time. Combining this with careful delineation of population boundaries using hundreds of genomes, we arrived at a startling conclusion: modern Prochlorococcus effective population sizes hover around 10 million, which are orders of magnitude smaller than the billions or trillions previously assumed. This means that genetic drift remains a potent force even in today’s oceans, shaping the fate of new mutations and maintaining slightly deleterious variants that selection would otherwise eliminate.
The final piece of the puzzle required us to reach back in time and directly constrain the population sizes of those ancient, ice-aged populations. Using agent-based models that simulate evolution under different population scenarios, we showed that the signature of radical mutation accumulation we observed could only arise if effective population sizes fell to 10,000 to 100,000 during the Snowball Earth. This is the same range as obligate endosymbiotic bacteria, organisms that are confined to host cells where Muller’s ratchet (the irreversible accumulation of deleterious mutations in small, non-recombining populations) is known to drive genome degradation. With reduced recombination imposed by physical isolation in refugia and critically small populations, the same ratchet likely clicked into gear for Prochlorococcus, randomly stripping away genes regardless of their utility.
Together, these findings tell a compelling story that spans scales from planetary ice ages to single nucleotide changes. The Prochlorococcus we see today, streamlined, abundant, and ecologically vital, carries the legacy of ancient bottlenecks that reshaped its genome through drift rather than design. This work challenges us to rethink how minimal genomes arise in the ocean and suggests that even the most successful modern lineages bear the scars of Earth’s dramatic past. By integrating deep-time geological events with cutting-edge evolutionary genomics, we are rewriting the rules of microbial evolution and opening new questions about how random chance, rather than optimal design, has shaped the invisible majority that powers our planet's biogeochemical cycles.
Key published / submitted papers in this area:
H. Luo. 2025. How big is big? The effective population size of marine bacteria. Annual Review of Marine Science (invited review article; 17:537-560)
H. Zhang, F.L. Hellweger, and H. Luo. 2024. Genome reduction occurred in early Prochlorococcus with an unusually low effective population size. The ISME Journal 18(1): wrad035
Z. Chen, X. Wang, Y. Song, Q. Zeng, Y. Zhang, and H. Luo. 2022. Prochlorococcus have low global mutation rate and small effective population size. Nature Ecology & Evolution 6(2): 183-194
H. Zhang, Y. Sun, Q. Zeng, S.A. Crowe, and H. Luo. 2021. Snowball Earth, population bottleneck and Prochlorococcus evolution. Proceedings of the Royal Society B 288(1963): 20211956
H. Luo, Y. Huang, R. Stepanauskas, and J. Tang. 2017. Excess of non-conservative amino acid changes in marine bacterioplankton lineages with reduced genomes. Nature Microbiology 2: 17091
Bacteria have driven Earth’s biogeochemical cycles for over three billion years, yet reconstructing when key lineages and metabolic innovations emerged remains fundamentally challenging. Unlike eukaryotes, bacteria left behind few diagnostic fossils. Our lab has developed a different approach: rather than searching for bacterial fossils that scarcely exist, we “borrow” temporal information from the abundant eukaryotic fossil record.
Three types of ancient endosymbiotic events create direct evolutionary links between bacteria and fossil-rich eukaryotes. Mitochondria evolved from within Alphaproteobacteria. Plastids evolved from within Cyanobacteria. And numerous bacterial symbionts have co-evolved with eukaryotic hosts across deep time. These connections allow us to propagate well-constrained eukaryotic fossil calibrations deep into the bacterial tree of life.
We first demonstrated this approach by showing that Rickettsiales originated nearly 700 million years before animals, challenging the long-held assumption that these intracellular pathogens co-diverged with their modern animal hosts. Protists, not animals, were the ancestral training grounds for these bacteria. We then extended the strategy to date ammonia-oxidizing bacteria, resolving debates about the order of nitrifier evolution and its relationship to the Great Oxidation Event. Critically, these analyses revealed that aerobic ammonia oxidizers (Gamma-AOB) predate anaerobic ammonia-oxidizing (anammox) bacteria, supporting a stepwise emergence of the nitrogen cycle where nitrite production by aerobes was prerequisite for anammox. Applying the same principles to flavobacteria, we linked their repeated marine-to-nonmarine transitions to the assembly and fragmentation of the Columbia supercontinent between 2.1 and 1.1 billion years ago, revealing how tectonic history shaped microbial diversification.
More recently, we formalized these ideas into a probabilistic framework (pRTC) that uses host-symbiont associations to constrain bacterial divergence times while accounting for uncertainty in ancestral lifestyles. This framework, applied across the bacterial tree of life, placed the last bacterial common ancestor at 4.0 to 3.5 billion years ago and established a robust timescale for testing hypotheses linking bacterial diversification to Earth’s geochemical evolution.
Parallel to these dating efforts, we developed machine learning tools that predict microbial phenotypes from genomic data. Our GBDT40-LR model identifies aerobic bacteria using just 40 conserved genes, most without direct roles in oxygen metabolism. Applied to over 80,000 genomes and integrated with our calibrated bacterial timetree, this placed the origin of aerobic bacteria at approximately 2.7 billion years ago, predating the Great Oxidation Event by several hundred million years and suggesting early life exploited localized oxygen oases before atmospheric oxygenation.
Key published / submitted papers in this area:
T. Liao, S. Chen, S. Wang, Y. Huang, S.K.W. Tsui, E.E. Stüeken, Q. Cao, and H. Luo. 2026. Non-canonical genetic markers resolve the pre-GOE emergence of aerobic bacteria in Earth’s history. Proc Natl Acad Sci USA 123(4): e2515709123
H. Zhang and H. Luo. 2025. The ecological diversification of flavobacteria: linking habitat transitions to Proterozoic supercontinent cycles. ISME Communications 5(1): ycaf210
S. Wang and H. Luo. 2025. Dating the bacterial tree of life based on ancient symbiosis. Systematic Biology 74(4):639-655
T. Liao, S. Wang, H. Zhang, E.E. Stüeken, and H. Luo. 2024. Dating ammonia-oxidizing bacteria with abundant eukaryotic fossils. Molecular Biology and Evolution 41(5): msae096
H. Zhang, S. Wang, T. Liao, S.A. Crowe, and H. Luo. 2023. Emergence of Prochlorococcus in the Tonian oceans and the initiation of Neoproterozoic oxygenation. (on bioRxiv)
T. Liao, S. Wang, E.E. Stüeken, and H. Luo. 2022. Phylogenomic evidence for the origin of obligately anaerobic Anammox bacteria around the Great Oxidation Event. Molecular Biology and Evolution 39(8): msac170
S. Wang and H. Luo. 2021. Dating Alphaproteobacteria evolution with eukaryotic fossils. Nature Communications 12: 3324
For decades, research on terrestrial nitrogen fixation has centered on symbiotic rhizobia that nodulate legumes. Our laboratory integrates evolutionary genomics, microbial ecology, and functional genetics to understand the origins and diversification of this critical trait, using the globally abundant genus Bradyrhizobium as a model system. Using this approach, we have shifted this focus by demonstrating that free-living nitrogen-fixing Bradyrhizobium represent the ancestral state from which symbiosis repeatedly evolved.
Through large-scale cultivation and genome sequencing of hundreds of novel strains from rice paddies, forests, and grasslands, we reconstructed the evolutionary history of nitrogen fixation genes. The earliest-diverging lineages are capable of free-living nitrogen fixation, establishing this as the ancestral condition. Nod factor-dependent symbiosis arose independently on at least three occasions via horizontal acquisition of symbiosis islands by distinct free-living ancestors.
This evolutionary trajectory is inscribed in genomic architecture. Free-living lineages maintain a highly conserved nif island (~50 kb) featuring the oxygen-protective gene glbO. Symbiotic lineages exhibit variable gene arrangements with universal glbO loss. Using genetic approaches, we validated that glbO protects nitrogenase from oxygen inactivation in fluctuating soil conditions but is dispensable within the microaerobic nodule environment.
Functional assays reveal a clear hierarchy: free-living strains exhibit robust nitrogenase activity under low oxygen, while symbiotic strains have largely lost this capacity outside the host. The nod-free but nodulating Bradyrhizobium phylogroup, enriched in rice rhizospheres, demonstrates exceptional nitrogenase activity and conserved nodulation ability with Aeschynomene legumes. These traits position them as promising candidates for sustainable agriculture through associations with non-legume crops.
Key published / submitted papers in this area:
L. Ling, S. Wang, J. Tao, M. Pervent, K.E. Ho, C. Sciallano, A. Camuel, N. Nouwen, E. Giraud, and H. Luo. 2026. The free-living wellspring of symbiotic nitrogen fixation in Bradyrhizobium. (under review)
L. Ling, A. Camuel, S. Wang, X. Wang, T. Liao, J. Tao, X. Lin, N. Nouwen, E. Giraud, and H. Luo. 2025. Correlating phylogenetic and functional diversity of the nod-free but nodulating Bradyrhizobium phylogroup. The ISME Journal 19(1): wraf030
J. Tao, S. Wang, T. Liao, and H. Luo. 2021. Evolutionary origin and ecological implication of a unique nif island in free-living Bradyrhizobium lineages. The ISME Journal 15(11): 3195-3206