Within cicadas such as those in the genus Tettigades, a world of microbes has evolved to provide nutrients to the cicadas. As reported in a pair of new studies at the University of Montana, researchers have discovered that the endosymbiont Hodgkinia cicadicola has split into at least two dozen lineages within cicada cells, in an apparent case of nonadaptive evolution. (Photo credit: Piotr Lukasik, Ph.D.)

By Viviane Callier

Since the origin of cicadas more than 100 million years ago, the bacteria Hodgkinia cicadicola has lived inside cicada cells, providing cicadas with amino acids and vitamins deficient in their diet of plant sap. In exchange, the cicada shelters the microbe and transmits it from one generation to the next through its eggs, ensuring its continued survival. It was a happy and mutually beneficial relationship for millions of years, but, in some lineages of cicadas, something strange happened about 4 million years ago: The endosymbiont started splitting inside the insect cells.

Viviane Callier

In fact, in some cicadas, the endosymbionts fragmented dozens of times, creating more than two dozen different lineages of microbes in the cicada cells, researchers at the University of Montana report in a pair of studies, published in November in Current Biology and December in Proceedings of the National Academy of Sciences (PNAS). In each cicada species, the genes from the ancient endosymbiont appear almost randomly distributed across the new endosymbiont lineages.

“The outcomes of the splitting process are really different in different cicadas,” says senior author John McCutcheon, Ph.D., associate professor at the University of Montana. That suggests that these speciation events are an example of nonadaptive evolution. “There’s nothing in this that suggests that the cicada is better off. … It’s happening due to random chance, and the cicada has to adapt to it,” McCutcheon says.

The discovery of the speciating endosymbionts was itself a result of random chance. In 2014, JT Van Leuven, McCutcheon’s student, sequenced a cicada expecting to find one Hodgkinia genome. But he found two, “which was weird. That’s not supposed to happen,” McCutcheon says. The researchers showed that the two Hodgkinia genomes resided in different endosymbiont cells, so the single endosymbiont lineage had split into two. The two new genomes appeared to have complementary sets of genes, and they required the presence of the other genome to function as a whole.

McCutcheon speculated that this phenomenon might be related to cicadas’ exceptionally long life cycle. He chose to study the genus Magicicada, the 13- and 17-year cicadas, because they have the longest life cycle. In their Current Biology paper, the researchers report endosymbiont lineage splitting in seven related Magicicada species.

The study in PNAS reveals genome splitting in a distantly related cicada group, the South American genus Tettigades. The researchers spent three field seasons collecting these cicadas in Chile and used extensive next-generation sequencing and microscopy to study the endosymbionts. They found that splitting happened at least six independent times in this genus. And this all started to happen between 4 million and 5 million years ago.

“In Chilean cicadas, you get the sense that something really important happened 4 million years ago,” McCutcheon says. That’s right around the time of the last Andean uplift, which had a big effect on the landscape. Forests became deserts and vice versa. “If you’re an animal that lives underground for many years and, when you emerge, all the trees are dead, you have a serious problem,” he says. Perhaps a cicada population bottleneck due to rapid ecological change led to the fixation of these splitting endosymbiont lineages. The researchers are currently investigating whether cicadas indeed experienced a population bottleneck at the time that would have left signatures in the cicada genome.

The speciating microbes created lots of problems for their hosts. First, the cicadas had to figure out how to package not one but dozens of endosymbionts in their eggs. Without the full complement of endosymbiont genes, the egg would die. That could require the cicada to make bigger eggs to fit all the microbes, potentially limiting the number of eggs they are able make—a hypothesis that the researchers currently are investigating. In addition, the different species of endosymbionts are not equally abundant (some are present in many copies, while others are rare) creating a gene dosage problem that the cicada had to solve. The researchers currently are using transcriptome data to understand how the cicada solves this gene dosage problem.

Viviane Callier is trained as an insect physiologist and is now a freelance science writer in Washington, DC. Twitter: @vcallier. Email: vcallier@gmail.com