Deep, deep in geologic time, some 600 million or 700 million years ago, the very first animals evolved on Earth. Their closest relatives that still live today include sponges, sea anemones and comb jellies. But exactly which of these is truly the closest relative to the very first animals has remained one of the most contentious questions in evolutionary biology. With few fossils of these early, squishy animals, their history has necessarily been muddy, and it has been challenging to reconstruct what happened.

A study published on May 17 in Nature resolves the relationships of these early animals by looking at the chromosomes of sponges, comb jellies, jellyfish and three close single-celled relatives of animals. By studying the pattern of chromosomes at the base of the animal evolutionary tree breaking and fusing together, a team of researchers at the University of California, Berkeley, University of Vienna, Monterey Bay Aquarium Research Institute and University of California, Santa Cruz, determined that comb jellies, more formally known as ctenophores, are in fact the closest relatives of the first animals.

“Understanding these deepest relationships in the animal tree of life is absolutely critical for reconstructing the history of the origin and evolution of a lot of the complex traits that we’re most interested in—things like the nervous system and animal symmetry,” says Casey Dunn, an evolutionary biologist at Yale University, who was not involved in the study.

The implicit assumption for more than 100 years was that the history of animal evolution was largely a stepwise addition of complex features in the animal lineage, Dunn explains. Chief among those widely held assumptions was that sponges are really primitive because they lack neurons and muscles. That led to the idea that they must have split off from the animal lineage before neurons and muscles originated. Comb jellies have muscles and a network of neurons, so they were thought to branch later.

But back in 2008, based on early information from the first sponge and ctenophore genomes, Dunn and his colleagues had proposed that comb jellies branched before sponges did. The researchers found that the inventory of these animals’ genes didn’t match the idea that sponges were a “snapshot of time before this machinery evolved,” Dunn says. Sponges already had genes that resembled those for neurotransmitters; perhaps these were used for cell-to-cell communication long before the evolution of neurons, with their specialized shape and function.

After that 2008 paper, dozens of studies appeared. Some were consistent with Dunn’s result, and some refuted it. “I personally have remained neutral on this debate,” says Paulyn Cartwright, an evolutionary biologist at the University of Kansas, “because applying subtly different models of evolution for how sequences evolve could change the result—meaning that the findings were not very robust one way or another.”

“So my conclusion was that it’s a very difficult problem,” adds Cartwright, who was not involved in the 2008 paper or the new study. “Part of the reason why it’s so challenging is because we’re looking at something that happened over half a billion years ago. And not only did it happen half a billion years ago, but it probably happened relatively quickly in geological time, so there’s not a lot of information to reconstruct these very ancient events.” Also, ctenophores have had half a billion years to undergo their own independent evolution, and they have a variety of features that are unique to their lineage.

In the Nature paper, the team took a new, creative approach to analyze the genomes of these early animals. Over hundreds of millions of years, gene sequences mutate so much that any signal about the relatedness of different lineages is washed out. “So you need something that evolves very slowly that you can track,” says Dan Rokhsar, an evolutionary genomicist at U.C. Berkeley, who oversaw the study. Instead of looking at alterations in nucleotides (single-letter changes in DNA), the method—developed by Rokhsar, along with Oleg Simakov and Darrin Schultz, both at the University of Vienna—focuses on larger-scale features in the genomes: groups of genes on chromosomes.

This technique is based on a simple idea: over evolutionary time, the order of genes on a chromosome gets shuffled via mutations—for instance, via inversions that flip the order of genes within a chromosome. Although their order may change, the genes on a chromosome form a kind of linkage group: they don’t usually shuffle with genes on other chromosomes. But on rare occasions, chromosomes can break and fuse, leading those linkage groups to mix. These events are rare enough that it is possible to trace them all the way back to the origins of the first animals.

The key insight is that chromosome fusion and mixing is as irreversible as the mixing of milk in a cup of tea. So the researchers deduced that if they observed fusion-with-mixing events that were shared between two lineages, then that event must have occurred in the common ancestor of those two lineages. The irreversibility of fusion-and-mixing events makes them particularly well-suited for resolving relationships in the animal tree that have resisted more conventional methods.

To elucidate the relationships at the base of the animal tree, the researchers assembled sequences of each chromosome for the comb jelly Bolinopsis microptera, two deep-sea sponges and three unicellular relatives of animals: a choanoflagellate, an ichthyosporean and a filasterean amoeba. They also used existing chromosome-scale genomes of cnidarians (sea anemones, jellyfish and corals, among others), sponges and amphioxus, or lancelet—an invertebrate that is very closely related to vertebrates and is a bilaterian, an animal with bilateral symmetry.

From this wealth of genomic data, the team discovered four fusion-and-mixing events shared by bilaterians (amphioxus), jellyfish, and sponges but not by ctenophores. If sponges branched before ctenophores, that would require these exact same four fusion and mixing events to have occurred independently in two lineages, the chance of which is vanishingly small. The researchers’ findings therefore provide strong support for the idea that ctenophores branched first. “This paper is a sea change in the discussion of these relationships and their evolutionary implications,” Dunn says.

“I’m very much convinced that [the researchers] have solved this debate because of the type of characters they’re using,” Cartwright says. “They have very strong data to support the early diverging ctenophores.”

What the finding means is that the ancestor of all animals, including sponges, already had a well-developed nervous system, and it probably was free-swimming, Cartwright adds. “We have to rethink the function and the structure of the early ancestor of animals. It wasn’t like a simple sponge, but it was likely something much more complex,” she says.

Another implication of the findings is that sponges lost a lot of the elements of a proper nervous system and muscular system because they’re filter feeders attached to the bottom of the ocean floor. The elements of a nervous system in the sponge genome may not be so much the beginnings of an animal nervous system as the remnants of a well-developed nervous system in the ancestor, Cartwright explains.

Rather than animal evolution proceeding as a gradual increase in complexity, it’s clear that evolutionary losses are part of the story. It has also become clear that early animals evolved unusual nerve cell features. Recent discoveries have shown that ctenophores have no synapses, the tiny connections between neurons. Instead the cells of their primitive nervous system, known as a nerve net, are fused together, forming a syncytium—“an entirely new way to build a nervous system,” Dunn says. And although sponges lack neurons, they have cells with neuronal features, called neuroid cells, in their digestive system.

One takeaway from this long quest is that, as more information is learned, researchers may find that early animal nervous systems are more diverse and innovative than we can currently imagine. Now we have a solid tree on which to pin them, providing a roadmap of sorts for future discoveries about the evolution of essential animal features.