Published November 12, 2013
Trevor Lawley keeps hundreds of samples of C. difficile in his freezer, each identified by the country in which the bacterium unleashed its unique brand of misery and death.
He tracked down Aus001 in Melbourne, Australia; collected Gla010 in Glasgow, Scotland; and picked up Lei017 in the Netherlands as part of an international hunt for the origin of “epidemic” C. difficile – a global menace that pumps toxins into the guts of its victims. It has spread around the world’s hospitals in the last decade, killing thousands.
Lawley, a Canadian with a flair for microbial forensics who now works at a leading British research centre, spent two years travelling the globe collecting hundreds of samples of C. difficile.
Then, in his lab at the Wellcome Trust Sanger Institute, near Cambridge, Lawley and his colleagues extracted the bacteria’s secrets.
Two strains of antibiotic resistant C. difficile that emerged in North America caused the global epidemic, the sleuths report.
One emerged in the northeast U.S. a decade ago; the second, which they call FQR2, surfaced in Quebec.
And it was FQR2 – the Quebec bug – that took off globally, becoming a scourge in the U.K., continental Europe and Australia, the team reports. “It was the biggy,” says Lawley.
He and his colleagues say the emergence of the potentially lethal pathogen was fuelled by antibiotics – wonder drugs that are increasingly the source of very big problems.
“They were the secret ingredient,” says Lawley, pointing specifically to fluoroquinolones, a class of antibiotics widely used in North American hospitals when the epidemic C. difficile strains appeared.
It is impossible to know exactly where FQR2 morphed into existence, but the intestines of a patient in a Quebec hospital is one scenario. Another is that it emerged unnoticed in the northeast U.S., then spread north.
It’s an odd fate for C. difficile, an ancient and formerly innocuous bacterium that can’t even grow in the presence of oxygen. It has co-existed with humans for eons and spends most of its time as dormant and almost indestructible spores. C. difficile can, however, come to life in the gut, and when conditions are right, it begins to multiply.
Doctors used to think of C. difficile and its spores, found in dirt, on food and animals, simply as a nuisance that could cause diarrhea.
“It was a low-grade problem,” says Dr. Mark Miller, former head of infectious diseases at the Jewish General Hospital in Montreal, who witnessed and helped document C. difficile’s transformation.
In 2000, doctors in Pittsburgh, Penn., began to see patients with severely inflamed colons. Eighteen patients died. An investigation pointed to a new virulent strain of C. difficile.
An eerily similar C. difficile surfaced in 2002 in Quebec. People undergoing hip replacements or heart bypasses started developing severe diarrhea and inflamed colons. Many ended up having their “toxic megacolons” removed, in a desperate bid to save their lives.
“We had never seen people getting so bad that you needed to take their colon out, or so bad that they had to go to intensive care unit, and certainly so bad that they would die of C. diff,” recalls Miller.
Miller and other infectious disease doctors in Montreal and Sherbrooke, anxious to figure out what they were dealing with, came up with a plan to track the cases and rates of complications.
“After three months, we knew we had a horrific problem,“ says Miller. “We knew it was a bad C. diff, a more virulent C. diff.”
Dr. Mark Miller does infectious diseases research at McGill University and the Jewish General Hospital in Montreal and is chief medical officer at bioMerieux, an infectious diseases diagnostic company.
They isolated and grew the microbes in the lab and soon realized the virulent C. difficile had also acquired the ability to live in the presence of fluoroquinolones, the potent antibiotic that doctors in Canada and the U.S. had begun heavily prescribing in the mid-1990s.
Fluoroquinolones, which prevent bacteria from replicating, were used to treat everything from urinary to respiratory infections. By 2002, they had become the most common antibiotic given to adults in the U.S. Almost one-quarter of all patients in one Quebec teaching hospital in 2003-4 were on fluoroquinolones.
Like many broad-spectrum antibiotics, fluoroquinolones disrupt the elaborate community of microbes living in people’s intestines by killing off beneficial as well as harmful bacteria.
But microbes with resistance, such as the new C. difficile, can continue to grow and multiply even when people take fluoroquinolones. And in many patients, the resistant C. difficile took over. Lawley likens it to the way weeds and shrubs move in after a forest is clear-cut.
Making the situation even worse, the resistant C. difficile could pump out extraordinary amounts of toxin: 16 to 23 times more toxin than garden variety C. difficile, according to one Quebec study.
This new “hypervirulent” C. difficile, as the doctors soon described it, can have a devastating impact, especially in people over age 60.
Once it takes hold in someone’s gut, the bacterium can invade the lining of the colon and pump out its extra-strength toxins with lethal impact. Cells in the colon start to disintegrate, and the colon begins to leak and ooze pus.
“You get these huge ulcers in your colon that can develop over hours to days,” says Miller. “It happens very quickly.”
There is also relentless diarrhea, with each episode releasing millions of C. difficile spores that can cling to toilets, bedpans and ambulances. The Quebec hospitals could not get rid of them.
The outbreak defied cleaning and infection control and the microbe moved from one hospital to the next. C. difficile spores, researchers later found, are so hardy, alcohol does not kill them. They can also be carried on shoes, clothes, people and animals, which helps explain how FQR2 skipped across continents.
Rod shaped C. difficile poking out of the “mucosal crypts” in the intestine of an infected mouse.
Miller, who has co-authored several studies characterizing the newly evolved C. difficile and the toll it’s taken, says more than 2,000 people died because of C. difficile infections during the two-year peak of the Quebec epidemic. “That’s conservative,” he says.
People started cancelling elective surgery and stayed away from the hospitals. “It was really quite a catastrophe,” says Miller.
The hypervirulent C. difficile, which Canadian microbe trackers classify as NAP1, was also on the move. It spread to Ontario hospitals and then across the country, infecting thousands of people in hospitals and nursing homes and causing an untold number of deaths.
Across the Atlantic, C. difficile also became a scourge, triggering unprecedented outbreaks in hospitals across the U.K. and Europe. The British government reports 7,916 deaths related to C. difficile at the peak of the epidemic in England in 2007, with about half of the death certificates describing C. difficile as the underlying cause.
But Lawley and his colleagues at the Sanger Institute, home to powerful genome sequencing machines, had the forensic tools to find out where NAP1, or “epidemic” C. difficile, emerged.
Lawley literally followed the trail of the NAP1’s “rapid transcontinental dissemination.” He collected samples of NAP1 from hospitals in the U.K., Europe, Asia, Australia and North America, amassing more than 300 strains isolated from infected patients.
Back in the lab, Lawley and his colleagues coaxed the bacteria to grow in oxygen-free cabinets, extracting enough DNA to sequence the organisms’ genomes and make a phylogeny – basically a family tree showing how the microbes are genetically related.
The analysis not only confirmed “epidemic” NAP1 C. difficile originated in North America but showed it had evolved twice. The C. difficile lineage, or strain, they call FQR1, first seen in Pittsburgh area in 2000, spread across the U.S. and later showed up in Switzerland and South Korea. “The second strain of C. difficile, FQR2, originated in Canada and spread rapidly over a much wider area, spreading throughout North America, Australia and Europe,” the Sanger Institute reports.
The analysis did not pinpoint where FQR2 originated, but Lawley says the bacterium likely emerged in either Quebec or the Northeast U.S.
And it show FQR2’s “descendants” were circulating in Birmingham, England, within months of the outbreak in Quebec and there were two other “trans-Atlantic transmission events” from North America into the U.K., two more into continental Europe and a more recent introduction into Australia. Lawley wouldn’t speculate on who or what carried the microbes across the continents.
Miller, who is now chief medical officer at bioMerieux, an infectious diseases diagnostic company, and who does research at McGill University and the Jewish General Hospital, describes the findings as “obviously fascinating.”
“Everyone has been pointing fingers at one another,” says Miller, referring to a long-running debate over whether the epidemic C. difficile should be called the Pittsburgh or Quebec strain. “In fact they are different.”
Michael Mulvey, who tracks C. difficile and other superbugs in Canada at the National Microbiology Laboratory in Winnipeg, says the findings shows the power of new genetic mapping tools that can detect tiny changes that occur when bacteria multiply and spread.
Mulvey’s team is taking a close look at 150 C. difficile samples collected by hospitals across Canada since 2004. They’re now feeding DNA from the organisms into their sequencing machines, which can spit out the genetic blueprint of a bacterium in less than 24 hours.
Electron microscope image of a colony of C. difficile growing in the lab.
The 100 bacteria analyzed so far are all related to the FQR2. “The same branch of the family tree as the Quebec strain,” says Mulvey.
Lawley says FQR1 and FQR2 appear to share a common ancestor, a virulent form of C. difficile that existed in nature. But it was when the virulent organism mutated and became resistant to fluoroquinolones antibiotics that the bacteria acquired the competitive advantage to take hold and thrive in hospitals, he says.
He says it’s not all surprising that the resistant, virulent C. difficile evolved twice, considering the way fluoroquinolones were so widely used. C. difficile needed just one small change in its genome to become resistant to the antibiotics. “So it was just a matter of time,” says Lawley.
The FQR1 and FQR2 strains continue to cause much misery, but the scientists say new C. difficile strains are now on the rise.
Mulvey says the hypervirulent NAP1 strain that was caused almost half the C. difficile infections in Canadian hospitals in 2008, is now down to about 30 per cent. Another strain, called NAP4, is on the rise and now causes almost 20 per cent of Canadian hospital infections.
It’s not known why NAP4 is increasing or where it came from. But Mulvey’s team is feeding the microbe’s DNA into the genome sequencing machines to try find out.
– Research for this story was funded in part by a journalism award from the Canadian Institutes of Health Research.
A microbe’s rap sheet:
Unlike health agencies in the U.S. and Britain, the Public Health Agency of Canada does not release comprehensive national data on C. difficile infections.
But reports from provincial and academic groups show the microbes continue to take a heavy toll. Onestudy released this fall estimates there were 37,900 C. difficile infections in Canada in 2012.
Ontario, Quebec and British Columbia have the highest rates of infection, and the numbers have been climbing in Alberta. Most people recover but C. difficile can cause relentless and fatal diarrhea, particularly in vulnerable older people in health care facilities.
Ontario had 42,421 cases of C. difficile infections between April 2002 and March 2010, according to a recent study. The province’s infection rate now runs at between 5,000 and 6,000 cases a year. Ontario does not release data on deaths associated with the C. difficile infections.
Quebec had 32,374 C. difficile infections in its hospitals in the eight years between 2004 and 2012, with 4,554 individuals dying within 30 days of infection, according to the province’s public health institute. The institute does not specify how many deaths are attributable to C. difficile infection and notes that many of the patients had serious underlying health conditions. Quebec hospitalsreported 3,748 C. difficile infections in 2011/12, the most recent data available.
British Columbia had 17,468 C. difficile cases between 2008 and March, 2013, according to the province’s infection control network, which starting releasing data in 2008. In the two years between 2008 and 2010, more than 1,000 individuals in B.C. died within 30 days of infection with C. difficile. The B.C. network no longer releases statistics on deaths associated with the C. difficile, saying the data is “open to misinterpretation.” B.C. had 3,246 C. difficile cases in 2012-2013, according to the most recentreport that shows how infection rates vary markedly in hospitals across the province.
Canada’s chief public health officer, Dr. David Butler-Jones, notes in his 2013 Report on the State of Public Health in Canada that the “mortality rates attributable” to C. difficile infection have more than tripled in Canada in the last 15 years. In 1997 there were 1.5 deaths for every 100 patients infected with C. difficile in Canadian hospitals participating in a national surveillance program, a rate that rose to 5.4 deaths for every 100 infected patients in 2010.