Cramped in a small submarine
2,500 meters below the Pacific’s surface in 2006, microbiologist Hiroyuki
Imachi scanned the ocean floor for signs of microbial life.

As the sub drifted over the
bottom of Japan’s Nankai Trough — a hotbed of understudied microbes living off methane
bubbling up from tectonic faults — Imachi spotted a nest of small clams against a
whitish microbial mat, suggestive of an active methane seep below. The submersible’s
robotic arm plunged a 25-centimeter tube into the blackish-gray sediment to
retrieve a core of muck.

It would take another 12
years of lab work for Imachi and colleagues to isolate a prize they hadn’t even
set out to find — a
single-celled microbe from an ancient lineage of Archaea, a domain of life superficially similar to bacteria. That
find could help biologists reconstruct one of life’s greatest leaps toward
complexity, from simple bacteria-like organisms to more complicated eukaryotes,
the enormous group of chromosome-carrying creatures that includes humans,
platypuses, fungi and many others.  

“Patience is very important
in doing successful science,” says Imachi, of the Japan Agency for Marine-Earth
Science and Technology in Yokosuka. He and his colleagues published their findings
in the Jan. 23 Nature, to
enthusiastic acclaim from fellow microbiologists. “I’m very lucky.”

Japanese research submarine
The Japanese research vessel Shinkai 6500 dove 2,500 meters into the Nankai Trough off of Japan’s Kii peninsula to sample the microbial diversity in the sediment around a methane seep in 2006.JAMSTEC (CC-BY-NC)

Many scientists think an
unusual meal kicked off the evolution of more complicated cells about 2 billion
years ago. An ancient archaean, the
theory goes, gobbled up a bacterium that, instead of being dinner, sparked a
symbiotic relationship in a process called endosymbiosis
(SN: 6/8/74). Eventually, the
bacterium evolved into mitochondria, the energy-producing cellular structures that
fueled the rise of complex life.

Living remnants of ancient
archaeal lineages persist in some of Earth’s most extreme environments, and
scientists are exploring these microbial hot spots for clues about the ancestor
of all eukaryotes. One such environment is the deep-sea floor. Despite making
up about 65 percent of Earth’s surface, biologists have only a faint picture of
the microbial multitudes that thrive there. Genetic sequencing of dredged up
mud has given biologists one way of studying these communities of bacteria and
archaea uniquely adapted to the cold, oxygen-less deep. But genes can reveal only
so much.

So scientists seek to grow
cultures of microbes in the lab to study what these organisms look like and how
they behave. But extreme microbes present unique challenges. Simply plating these
organisms on a petri dish, providing nutrients and waiting for growth hadn’t
ever worked —
possibly because scientists weren’t effectively re-creating the microbes’
extreme environment, says Masaru Nobu, a microbiologist at the National
Institute of Advanced Industrial Science and Technology in Tsukuba, Japan, who
joined Imachi’s project after it started.

So Imachi, Nobu and their
colleagues tried to re-create a methane seep in the lab, drawing inspiration
from a bioreactor used to treat municipal sewage. The team pumped methane gas
into a meter-tall cylindrical chamber, kept at 10° Celsius and stacked with polyurethane sponges that mimic
porous deep-sea sediment. A slow, steady flow of artificial seawater kept the
sponges saturated.

The team then watered down a
clump of mud from the Nankai Trough sediment core, sopped up the slurry with
the sponges, stacked them in the reactor — and waited.

“There was a lot of
nervousness,” Nobu says of that time in December 2006. “We didn’t know if we’d
get what we wanted.”

Masaru Nobu (and Hiroyuki Imachi
Microbiologists Masaru Nobu (left) and Hiroyuki Imachi pose in front of the bioreactor that they used to grow microbes from sediment dredged up from the deep sea.M.K. Nobu and H. Imachi

Each year, the researchers
sequenced genes of microbes in the sponges. After a volatile first couple of years,
the microbial community began to stabilize and grow. “Most of the organisms
that were active in the reactor were organisms that were actually active in the
natural environment,” Nobu says. With a stable community of thousands, if not
tens of thousands, of different kinds of microbes to draw on, the team could try
to pick out and grow individual strains.

Samples from the reactor
were placed into 200 glass bottles, each filled with a different energy source
and cocktail of antibiotics to weed out bacteria and allow different archaea to

The team had its first
eureka moment in 2011, detecting an archaean
new to science that they called MK-D1 in very low numbers amid numerous bacterial
strains in one bottle. But each time the team tried to isolate the archaean in a new bottle, the microbe just wouldn’t grow. Months
of trial and error followed. “It was really frustrating,” Nobu says.

Then, the researchers had an
idea: Perhaps the microbe was actually growing, but at a slow pace, a result of
its deep-sea home. “It’s very cold down there, there’s not a lot of energy,”
Nobu says.

So the team measured growth
with a more sensitive technique, called quantitative PCR, that can quantify
abundance from whiffs of DNA. Sure enough, MK-D1 was there and growing, just
more slowly than any other single-celled microbe ever cultured. E. coli, for instance, can replicate itself
in about 20 minutes. MK-D1 takes two to three weeks to divide.

“No microbe we knew about
grew this slowly,” Nobu says. “Understanding this was a revelation.”

Meanwhile, another archaea
discovery in 2015 rocked the world of microbial ecology. A new group dubbed
Asgard archaea had been discovered from genetic material dredged up from a
hydrothermal vent in the Arctic Ocean. Asgards have many eukaryotic genes, leading
some scientists to argue that Asgards are the closest living relatives of ancient archaea that may have given rise to all
complex life on Earth (SN: 12/15/15).

Sign Up For the Latest from Science News

Headlines and summaries of the latest Science News articles, delivered to your inbox

Imachi and Nobu were stunned
when DNA evidence confirmed that they’d unwittingly spent the last nine years
cultivating their own Asgard, MK-D1. If it could be isolated, Imachi’s team
would be the first to actually glimpse a living member of this exciting but mysterious

The researchers finally got a
stable culture of MK-D1 to thrive — with a bacterial partner that it needs to
survive — and in 2018, took their first look under a microscope. The neat, tiny
spheres seen at first seemed unlikely to be the sort of thing that may have begotten
complexity. But over months, the microbes grew odd, tentacle-like protrusions. Imachi
“initially thought the sample had been contaminated,” he says. But the
observation was sound, prompting the researchers to propose a model for how
these tentacles might have ensnared other microbes — a probable first step in endosymbiosis.

The team gave MK-D1 a proper
name, Prometheoarchaeum syntrophicum,
after the Greek god Prometheus who, the myths say, introduced fire to humanity.
Much remains to be learned about P.
and what, if anything, it can tell us about our origins. In
the meantime, Imachi is still sifting through the microbes in his reactor.

As he puts it, “uncultured
microbes are waiting.”


    Related Articles