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A report in Science 2 years ago that linked a mouse retrovirus, XMRV, to chronic fatigue

syndrome astonished scientists and patients alike. But the theory soon began to take hits,

and now, to all but a few researchers, it has completely unraveled

NEWSFOCUS

Done. Case closed. Finito, lights off, The End.

For the past 2 years, a controversy has roiled around the purported

link between a mouse retrovirus, XMRV, and chronic fatigue syndrome

(CFS), a baffl ing, debilitating disease with no known origin. Many

researchers who have followed this saga closely thought that a defi nitive

study, published online this week by Science (http://scim.ag/xmrv-cfs)

and conducted by nine labs—including the main proponents of the the-

sis—would fi nally bring a halt to the impassioned debate.

Think again.

The uproar began with an October 2009 paper in Science that found

XMRV in the blood of two-thirds of the CFS patients examined. A

steady assault on the report soon began, with more than a dozen labs

failing to replicate it to date and several asserting that contamination

must have occurred. The leader of the team that conducted the study,

Judy Mikovits of the Whittemore Peterson Institute for Neuro-Immune

Disease (WPI) in Reno, Nevada, resolutely maintained that her lab had

no evidence of contamination and that it could repeatedly fi nd the virus

with its techniques. Millions of dollars have gone into clarifying the

question, which has had far-reaching consequences for people with CFS

and, if the virus lurked in the blood supply, the public at large.

The study just published found that none of the nine labs could repro-

ducibly detect XMRV or relatives of the virus in blood samples distrib-

uted under a blinded code. Pounding another nail into the coffi n, Science

is also running a partial retraction (http://scim.ag/R-H-S) of the origi-

nal paper, as a contributing lab found that it in fact had a contamination.

In an unexpected twist to this operatic saga, Mikovits co-authored

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the Science Express paper and has no quarrel with the results. Her col-

laborator, Francis Ruscetti, a retrovirologist at the U.S. National Cancer

Institute (NCI) in Frederick, Maryland, who is a co-author of both the

original Science report and the new one, concurs. “Where there is dis-

agreement is in the interpretation,” Ruscetti says.

By their lights, the new study—conducted by the Blood XMRV

Scientifi c Research Working Group sponsored by the U.S. National

Heart, Lung and Blood Institute—does not rule out the possibility

that mouse retroviruses infect people with CFS. “The conclusion of

the Blood Working Group was that

we don’t have a reproducible

assay to detect XMRVs in the

blood—not that they weren’t

in the patients at all,” Mikov-

its says. Ruscetti adds that the

working group analyzed their

original patients but used samples

taken a few years later.

If this seems like wordsmith-

ing and splitting hairs, welcome to

the confusing, maddening world

of XMRV. Mikovits and Ruscetti,

who have become increasingly

isolated from the broader scien-

tific community, now say their

original paper erred by focusing

on a single XMRV isolate that

turned out to be a contaminant.

They say that isolate is but one

of many XMRVs, which belong

to a still larger family of gamma-

retroviruses. They also contend

that the virus may lurk in tissues,

only traveling to the blood occa-

sionally. “We still stand by our

data that we isolated gamma-

retroviruses from patients with

CFS and also from healthy con-

trols,” says Mikovits, who has taken a more public role than Ruscetti in

battling critics and reaching out to supporters.

Mikovits has become something of a savior in the community of

people with CFS (also known as myalgic encephalomyelitis, or ME),

who for decades have endured charges that the disease is psycho-

somatic. The 2009 Science paper shouted out that CFS may well

have a clear biological cause, and, in turn, raised hopes of effec-

tive treatments and even a cure. The new

fi ndings give her “great pause,” yet she sus-

pects they’re but a speed bump. “I haven’t

changed my thinking at all,” she says. And

she worries that the Blood Working Group

conclusions will confuse people with CFS,

some of whom got wind of the results early

in the blogosphere and contacted her in a panic. “I had 15 suicidal

patients call me last week,” she says.

In scientifi c circles, Mikovits has developed a less fl attering reputa-

tion. Critics have accused her and her backers of stubbornly wedding

themselves to a thesis and moving the goalposts with each study that

challenges their conclusions. Even disease advocates who welcome the

attention XMRV has brought to CFS believe the time has come to put

this line of research to rest. “It’s hard to say that this has not received a

fair appraisal,” says Kimberly McCleary, president of the CFIDS Asso-

ciation of America, a patient group in Charlotte, North Carolina.

Many retrovirologists wish the entire controversy, which has

ensnarled dozens of labs and cost millions of dollars, would simply

disappear. “All of it’s a waste of money and it’s wrong,” says Robert

Gallo, head of the Institute of Human Virology in Baltimore, Mary-

land. “It’s like a bad dream.”

XMRV owes its discovery to a little-known enzyme called

RNase L that helps the body battle viruses—and has ties to both

prostate cancer and CFS.

In the early 1990s, a few CFS

researchers reported that their

patients had higher levels of RNase

L than healthy people had, sug-

gesting that this natural viricide

signaled an undiagnosed infec-

tion causing the disease. RNase L

levels, they contended, provided a

long-sought “biomarker” for CFS,

which has caused much confusion

because clinicians use varying cri-

teria to make a diagnosis. It also

could potentially help researchers

gauge the effectiveness of treat-

ments, as a drop in the enzyme

would mark a decrease in under-

lying, undetected infections. This

idea became central to tests using

Ampligen, an immune modulator

made by Hemispherx Biopharma

in Philadelphia, Pennsylvania, to

treat CFS.

One of the clinicians testing

Ampligen was Daniel Peterson

of Incline Village, a town on the

northern shore of Lake Tahoe,

the popular tourist destination on

Nevada’s border with California.

Peterson had investigated a mysterious outbreak of CFS that began in

Incline Village in 1984 and had become one of the world’s best-known

CFS clinicians. His work drew the attention of Harvey Whittemore, a

prominent Reno attorney who lobbied on behalf of the gaming industry

and later became a major real estate developer, and his wife, Annette, a

teacher of children with cognitive disorders. The Whittemores’ daugh-

ter, Andrea, had developed CFS at age 12, and at Peterson’s suggestion

she began receiving Ampligen.

With the Whittemores’ fi nancial support, Kenny De Meirleir, a

Belgian physician who had an RNase testing lab in his own country,

helped them open RedLabs USA in Reno to monitor the enzyme in

U.S. patients taking Ampligen. Vincent Lombardi, who would soon

play a prominent role in the XMRV saga, co-founded the new lab and

served as its director of operations.

Lombardi was a late bloomer in science. In 1990, while he worked

as a securities trader in Lake Tahoe, he decided to pursue an under-

graduate degree at a local school. As part of a biostatistics class, he

worked with Peterson, analyzing immune parameters of CFS patients.

After his graduation in 1995, he ran investment companies and also

worked as a marketing director for a pawnbroker business. He went

on to pursue a Ph.D., fi rst studying RNase L but switching to peptides

in the tobacco hornworm, which he completed at the University of

Nevada, Reno, 2 years after starting RedLabs USA.

Onlinesciencemag.org

Podcast interview

with co-author

Jon Cohen.

“I don’t care if

nobody else in

the world wants

to work on it.

Fine, leave us

alone!”—JUDY MIKOVITS,

WHITTEMORE PETERSON

INSTITUTE

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In 2002, meanwhile, a team of prostate cancer researchers had discovered that a muta-tion in the RNase L gene frequently occurs in families in which men are prone to the early-onset form of that disease. A few years later, one of those investigators, Robert Silverman of the Cleveland Clinic in Ohio, began probing for a viral link: If RNase L fi ghts viruses, he reasoned, then perhaps having a crippled form of the enzyme opened the door for a cancer-causing infection.

Silverman teamed up with Joseph DeRisi and Don Ganem, two veteran virus hunters at the University of California, San Francisco (UCSF), who had developed a microarray called ViroChip that had proved its mettle in identifying new viruses. The assay con-firmed Silverman’s hunch: In the prostate cancer tumors, ViroChip found a novel retro-virus, apparently a cousin of a known mouse virus and therefore given the ungainly name of xenotropic murine leukemia virus–related virus (XMRV).

Silverman fi rst reported the existence of XMRV and its prostate cancer link in April 2005 at an HIV meeting in a mountain resort in Banff, Canada. “My talk was well received, people were clearly interested, but it did not receive that much attention,” Silverman wrote to Science in an e-mail delivered through a public affairs manager—the only form of communication the Cleveland Clinic would allow. (XMRV has become such a radioactive topic that several institutions restricted what their researchers could discuss, and both Ganem and DeRisi declined to talk to Science about their XMRV work.)

In March 2006, Silverman, DeRisi, Ganem, and colleagues pub-lished their report in PLoS Pathogens, a respected but decidedly low-key choice for the description of a new retrovirus that appeared to infect humans—the only three others are HIV and HTLV-I and -II—and had ties to a serious disease. Although they wrote that there was a “strong association” between XMRV and the RNase L mutation, the researchers still had misgivings about whether the virus contributed to prostate cancer.

In a commentary in the 30 January 2007 issue of the Proceedings of

the National Academy of Sciences (PNAS), mouse retrovirologist Hung Fan of UC Irvine called it an “exciting” discovery. But others were skeptical. Gallo, whose lab played a central role in the discovery of all three known human retroviruses, contacted PNAS’s editor to com-plain. The commentary was “substantially over the top,” says Gallo, who saw no compelling evidence that this supposedly new human retrovirus caused disease and doubted that a mouse retrovirus could even infect humans. “Once claims of etiology were made, I just gasped for breath,” Gallo says. “My own experience argued to me that it’s best to stay away from this one.”

Around the time Silverman fi rst publicly described XMRV, Annette Whittemore took on one of the most ambitious pursuits of her

career: using her family’s wealth and powerful contacts to build a full-fl edged research and clinical institute devoted to CFS. Peterson would head a medical research team to elucidate the causes of the disease and provide state-of-the-art care. Nevada Governor Kenny Guinn and the current U.S. Senate Majority Leader, Harry Reid (D–NV)—both fam-ily friends of the Whittemores—helped arrange government funding,

while the Whittemores’ alma mater, the University of Nevada, Reno, offered about 1400 square meters of lab space within a new, gleam-ing $78 million Center for Molecular Medicine. It would take 4 years for that building to open, so the nascent Whittemore Peterson Insti-tute (WPI) set up shop in borrowed space on campus in 2006. Annette, WPI’s president, hired Mikovits to run the lab.

Mikovits had come into the CFS world by a circuitous route. She had spent more than 20 years at NCI, fi rst as a technician studying HTLV-I and HIV with Ruscetti, who served as her Ph.D. adviser. In 2001, she married and moved to southern California, becoming chief scientifi c offi cer at EpiGenX Biosciences in Santa Barbara, which aimed to use a new epigenetic approach to develop drugs and diagnostics.

In 2006, Mikovits became a consultant to a CFS-related nonprofi t foundation that Annette Whittemore co-founded, which explored the link between the disease and another virus, human herpesvirus 6 (HHV-6), that had been discovered in Gallo’s lab. At a meeting in Bar-celona, Spain, that year, Mikovits spoke to Whittemore for the fi rst time and heard Peterson give a talk. Peterson focused on a non-Hodgkin’s lymphoma in some of his CFS patients, and Mikovits smelled a virus. She offered to work with him, and Whittemore helped set up a collabo-ration. Later that year, Mikovits joined WPI.

She soon enlisted Ruscetti, who had worked in Gallo’s lab when it discovered HTLV-I, to screen blood samples from Peterson’s patients for viruses. Intrigued by the RNase L link to XMRV, Mikovits and Lombardi—who by then had joined WPI as well—met Silverman in October 2007 at a prostate cancer conference in Lake Tahoe, where they discussed the possible role of XMRV in CFS. Silverman was happy to collaborate and sent WPI a clone of the virus, known as VP62. The institute could use it as a reference to start hunting for the virus in CFS patient blood samples that Peterson had stored.

A little over a year later, on 18 November 2008, Mikovits had the fi rst evidence of XMRV in CFS patients. Working with Ruscetti and Silverman, the group amassed evidence that the virus occurred in 67% of 101 CFS patients and 3.7% of 218 healthy controls. If the latter number was representative of the general population, up to 10 million

“I began comparing

Judy Mikovits to Joan

of Arc. The scientists

will burn her at the

stake, but her faithful

following will have

her canonized.”—JOHN COFFIN,

TUFTS UNIVERSITY

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healthy people in the United States alone might

be infected, and the virus might be spreading

through blood donations and organ transplan-

tation—a silent epidemic of frightening pro-

portions. Mikovits, Ruscetti, Silverman, and

their co-authors submitted a paper to Science

in May 2009. The paper, of which Lombardi

was first author, did not claim that XMRV

caused CFS, noting that the disease might sim-

ply make patients more vulnerable to infection.

Causality “is probable but not defi nitive at this

time,” Lombardi et al. stated.

But what they were asserting was stunning

enough: WPI, NCI, and the Cleveland Clinic

had all found evidence of XMRV in the same

CFS patients. Some of these people had par-

ticipated in a 2007 NCI drug study, too, and

their blood samples, stored separately, also

tested positive, “ruling out the possibility of

lab contamination as a source,” the authors

wrote. When the team calculated the so-called

p-value—which generally needs to be under

0.05 for a fi nding to be considered signifi -

cant—they arrived at the astonishingly

low number of 8.1 × 10–35.

Oddly, Peterson, who had supplied

the patient samples, was not one of the

authors. His name was left off—and he was

kept out of the loop on the study’s results—

because of worries that he might prematurely

tell his patients, Mikovits says.

The manuscript didn’t convince Science.

After reviews by three referees and

members of the Board of Reviewing Editors

(provided for this article by Mikovits), the

editors rejected the paper. “Although the ref-

erees were intrigued by your fi ndings, they

had a number of serious reservations,” read a 4 June letter, which

included excerpts from reviewers.

The rejection letter noted that Science would re-review the paper

if the authors could both retain the “novelty of its main message” and

“address the referees’ concerns with new data rather than with counter-

arguments.” But the criticisms were substantial. “Chronic Fatigue Syn-

drome is full of false alarms,” wrote one advisory board member, “and

the detection of XMRV could be false positive PCR.” An otherwise

enthusiastic referee wrote that the “one major caveat I have is that the

issue of potential contamination has not been completely dealt with.” A

second referee found it odd that the genetic sequence of XMRV derived

from CFS patients and the virus earlier discovered in prostate cancer

were 99% similar. This “seems very unlikely and may indicate con-

tamination, despite the evidence presented to the contrary,” the referee

warned. One also wondered why they omitted Peterson as a co-author.

Mikovits and her co-workers addressed many of the critiques, and

on 14 July, they resubmitted the paper, this time with Peterson’s name

on it. The next week, a committee—which included John Coffi n, a

prominent retrovirologist at Tufts Sackler School of Graduate Biomed-

ical Sciences in Boston, and Silverman—organized a workshop at NCI

with Mikovits and Ruscetti to help the institute better understand the

unanswered scientifi c questions and the potential public health ramifi -

cations. Dusty Miller, a retrovirologist at the University of Washington,

Seattle, who had studied XMRV, left the meet-

ing convinced that the link to CFS was real.

The group had not only detected the virus using

PCR but also grown it from patients’ cells and

found antibodies to it. “It sounded really good

to me because they had all these different lines

of evidence,” Miller says. The fact that an NCI

veteran such as Ruscetti endorsed it helped

convince him. “Frank said, ‘I grew it with my

own hands,’ ” Miller recalls. “At the time, it

sounded really exciting.”

Coffi n, who chaired the meeting, thought

Mikovits and her colleagues had persuasive

evidence, especially because XMRV formed

its own branch on a genetic family tree of

mouse retroviruses, strengthening the case that

it was not a contaminant.

The next day, as Mikovits sat at Washing-

ton, D.C.’s Dulles International Airport wait-

ing for her fl ight back home, she received a

phone call from Science, indicating for the fi rst

time serious interest in publishing the paper.

Science accepted the Lombardi paper on

31 August and published it online on 8 October

2009, accompanied by a supportive commen-

tary that Coffi n co-authored. “There are several

lines of evidence that transmission happened

in the outside world and was not a laboratory

contaminant,” the commentary declared. The

publication catapulted Mikovits into the spot-

light, where she has remained ever since.

“Here we go again.” That’s what

crossed Kimberly McCleary’s

mind when she read the fi rst headlines

about the paper that week in her offi ce

in Charlotte. In her 20-plus years at

CFIDS, McCleary had seen many infec-

tious agents fi ngered as a potential cause of

CFS. HHV-6. Epstein-Barr virus. Mycoplasma. Adenovirus. Cyto-

megalovirus. HTLV-I and -II. And on and on. Each time, hopes were

dashed as scientists closely evaluated the suspects.

The string of disappointments had made McCleary cautious,

and CFIDS urged patients not to get carried away. “We tried to tem-

per things early on, and we were criticized heavily for raining on the

parade,” she says. Still, many patients went wild with joy. “ME/CFS

patients have never seen anything like this,” wrote Cort Johnson in an

item, “Game Changer,” posted on his popular blog Phoenix Rising.

Lombardi et al. had made XMRV a superstar. “Here was this myste-

rious disease (CFS-ME) with no known cause, all of these patients suf-

fering, and along comes a new virus that associates with the disease,”

Silverman says. “It seemed to some at the time to have the makings of

a medical breakthrough.”

But many scientists were skeptical. Simon Wessely, a psychiatric

researcher at King’s College London who has long studied CFS, says

the virology went over his head, but the fact that fully two-thirds of CFS

patients harbored the virus was an alarm bell. CFS, whose defi nition

has been the subject of years of debate, is far too heterogeneous a phe-

nomenon for that, he says.

With a few U.K. colleagues, Wessely started penning a letter to

Science to address what they saw as methodological fl aws in the paper.

“Here was this mysterious disease, …

and along comes a new virus. … It seemed to …

have the makings of a medical

breakthrough.”—ROBERT SILVERMAN,

CLEVELAND CLINIC

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One problem was the unclear patient selection, they wrote. If all patients came from Nevada and healthy controls were from elsewhere, then per-haps XMRV wasn’t a CFS-related virus, Wessely says, but something that happened to be more prevalent in the gambling state.

Wessely also contacted Jonathan Weber, a retrovirologist at Imperial College London, to set up a study of British CFS patients. “Our reac-tion was, ‘It’s probably wrong, but if it’s true, it’s a pretty big advance. So it’s worth testing,’ ” he says.

The fi rst published critique appeared a few weeks later—and it would prove to be prophetic. On 18 November, Patrick Moore, a cancer virologist at the University of Pittsburgh in Pennsylvania, and his post-doc Masahiro Shuda published a blistering commentary on Faculty of 1000, a Web site that evaluates reports in what it calls postpublication peer review.

“Unfortunately, in my field, there’s a tendency with any new virus to hope that it is causing a disease,” says Moore, who co-discovered HHV-8 and helped prove that it causes Kaposi’s sarcoma. “One has to be so

cautious about that. A lot of stories sound good, but they’re built on a house of cards.”

As Moore and Shuda noted, Mikov-its’s group used a technique called “nested PCR” to identify XMRV infections, which, they wrote, “is inherently prone to inter-mittent false positivity that has occurred in our lab and many others.” What’s more, the researchers had not randomized and blinded patient and control samples, a standard way to protect against bias and to detect errors. Together, Moore and Shuda wrote, this was “a recipe for uncontrolled PCR contamination.”

The next month, a study co-authored by Silverman and published online by Virology showed that AZT, an anti-HIV drug, worked against XMRV in test tubes. Again, CFS patients were overjoyed. Not only had a likely culprit of their suffering been found, drugs already on the market might treat it.

Of course, no one had proved that the virus actually caused CFS. But by the start of 2010, three commercial labs, one of which was WPI’s VIP Dx (formerly RedLabs USA), offered an XMRV test.

Peterson, sitting in one of his practice’s waiting rooms in Incline Village in June, explained that his patients split into two groups. One had ridden the CFS roller coaster so many times that these patients wanted validation and replication before buying the XMRV story. “There’s another group that, for whatever reason, has made this ill-ness into a religion and becomes polarized into who believes and who doesn’t believe,” he said. Increasing numbers of believers began taking XMRV tests, which cost $500 or more, and, if they were positive, ask-ing their physicians to prescribe anti-HIV drugs, says Peterson, who by then had severed ties with WPI over a contractual issue.

When a Science reporter visited WPI the same month, Annette Whittemore and Lombardi both strongly defended the decision to sell an XMRV test before evidence existed of a causal link. (The group has fi led related patent applications, too.) “Every physician who requests patient testing is aware of contamination and the potential of false

hope,” Lombardi said. “We’re not the doctors ordering these tests for our patients.” Whittemore stressed that all profi ts would go back into the institute. “This wasn’t set up to make money, and it never has,” she said. Whittemore also dismissed critics—a group that included Peter-son—who cringed at CFS patients taking antiretrovirals. “How many years does this patient population have to be impacted and their lives destroyed?” she asked.

Andrea Whittemore was one of the CFS patients who tested posi-tive for XMRV.

The flood of negative data started in January 2010, when Wessely, Weber, and their colleagues reported in PLoS ONE that

they couldn’t fi nd any trace of XMRV in 186 British CFS patients. By the end of February, two more negative reports were published, one of them by Jonathan Stoye, Coffi n’s co-author on the favorable Science commentary that accompanied the Lombardi paper.

Back in Reno, Mikovits and Lombardi began feeling besieged. “After the fi rst neg-ative study, it was a dog pile,” Lombardi says. “Let’s be honest: A number of peo-ple in the mainstream medical community heard chronic fatigue, and they rolled their eyes and laughed.”

The problem, as they saw it, was that nobody was following their recipe exactly. Many labs failed to fi nd XMRV using PCR because the virus exists in scant amounts, they said—so much so that a patient can test positive on one bleed and negative on the next. Mikovits said they overcame this problem by fi rst mixing patient blood samples with an uninfected cell line that is especially permissive to the virus and coculturing for 8 weeks. Other researchers, including XMRV discoverer Silverman, questioned the need for this step.

Seven months after Lombardi et al., Science published Wessely’s critique and

two others, which discussed patient selection problems and the growing list of negative studies, including three

that failed to find XMRV in prostate cancer. None of the Technical Comments,

however, mentioned the possibility of contamination. Despite a vigorous defense by Mikovits and Ruscetti in the same

issue of Science, the three comments further eroded confi dence in the fi ndings. But then, 2 weeks later, a research group unconnected to WPI would rekindle the fading hope that the link between XMRV and CFS was real. It would also spark a peculiar new controversy.

That study originated with Shyh-Ching Lo, an infectious-disease researcher at the U.S. Food and Drug Administration (FDA) who

took a keen interest in CFS. In the early 1990s, Lo had tested a theory that a microbe called Mycoplasma was involved in CFS. For that study, he had received blood samples from Anthony Komaroff, a physician who had treated hundreds of CFS patients at Brigham and Women’s Hospital in Boston. Like so many others, the lead had proved spurious.

After XMRV surfaced as a new candidate, Lo suggested to Komaroff they test the same patient samples, stored at –80°C and

“We tried to temper things early on, and we

were criticized heavily for raining on the parade.”

—KIMBERLY MCCLEARY,

THE CFIDS ASSOCIATION OF AMERICA

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untouched for almost 2 decades. Komaroff was enthusiastic. Lo also

contacted Harvey Alter, an infectious-disease specialist who works on

transfusion medicine at the U.S. National Institutes of Health’s Clinical

Center in Bethesda, Maryland. Alter—who won the prestigious Lasker

Award for his role in the discovery of hepatitis C—provided Lo with

blood samples from healthy people to serve as controls.

It was Alter who accidentally dropped the bombshell news. At a

blood bank meeting in the Croatian capital Zagreb in late May 2010,

he presented an overview of current blood safety issues, and his Power-

Point presentation had one slide on what he called the “Agent du jour,”

XMRV. Although Alter didn’t present any evidence, the slide was blunt:

“The data in the Lombardi et al. Science manuscript are extremely

strong and likely true, despite the controversy,” one bullet point said.

“We (FDA & NIH) have independently confi rmed the Lombardi group

fi ndings,” read another.

After the editor of a Dutch newsletter discovered the slide on

the meeting’s Web site and sent out a press release, the CFS world

exploded, again. “Is this it, then, the big one???? Holy ****!!!!!” one

blogger wrote on the Phoenix Rising forum.

Some researchers had doubts. True, Alter had a stellar reputa-

tion, but he wasn’t a retrovirologist. And Lo in the 1990s had pushed

the widely discredited theory that a Mycoplasma infection played an

important role in HIV causing AIDS.

Yet news of the impending confi rmation had a big impact on a spe-

cial XMRV panel at the AABB, an international association focused on

transfusion medicine. The panel, which had heard about the Lo-Alter

study before the news broke, recommended on 18 June that AABB

members discourage CFS patients from donating blood.

Patients and scientists alike were eager to see the data. Lo and Alter

had written a paper accepted by PNAS, but they ran headfi rst into an

institutional roadblock: Retrovirologists at the U.S. Centers for Disease

Control and Prevention (CDC) had their own paper in preparation that

showed no evidence of XMRV antibodies, proteins, or DNA in well-

characterized CFS patients. Faced with contradictory results from two

teams of researchers under its purview, the U.S. Department of Health

and Human Services (HHS) ordered the groups to delay publishing

until each could review the other’s data.

Patients cried foul. Many had long distrusted the federal govern-

ment, and especially CDC, for not taking CFS seriously and suppress-

ing research results. Now, they said, CDC was trying to bury another

theory it didn’t like. The author of a blog, CFS Central, called for

aggressive, ACT UP–style pro-

tests at blood banks to demand

publication: “I believe we need

to act quickly, before the FDA/

NIH paper is killed.” Some scien-

tists thought the federally ordered

delay impinged on scientific

freedom. Why not publish the

competing papers and let other

researchers scrutinize them? “It

was very strange business,” Cof-

fi n says. Alter says nothing sinis-

ter was afoot and that the papers

were never in peril. Retrovirology

published the CDC study on

1 July 2010, and the Lo-

Alter study ran in PNAS on

23 August.

Once they saw it, many

researchers concluded that

the Lo-Alter data didn’t confi rm the Lombardi paper at all. They had

found that 86.5% of samples from CFS patients harbored DNA from

mouse retroviruses as did 6.8% of healthy controls, but it was not

XMRV; the sequences were more closely related to a different, well-

known group, the murine leukemia viruses (MLVs). It was as if a new

suspect suddenly had been nabbed. Miller says the fi ndings actually

argued against Lombardi et al. “If XMRV is everywhere and these

guys are doing the same experiment, why didn’t they fi nd XMRV?”

he asks. Alter now says he regrets asserting that the paper confi rmed

the XMRV results and that it was “naïve” to show the slide in Zagreb.

“That is something I shouldn’t have said because we really hadn’t

found it,” he says. “I somehow got to be the spokesperson for this. I

had no idea what it would mean.” Lo agrees that the team didn’t fi nd

the virus that Mikovits and Lombardi had reported.

Mikovits to this day contends that the Lo-Alter paper confi rms

Lombardi et al. and insists that from the beginning, she viewed XMRV

as one of many gammaretroviruses, which includes the MLVs, involved

with CFS. In the Lombardi study, some patients tested negative in PCR

tests for XMRV and yet produced MLV-related proteins, she claims,

but they decided to count them as negatives. She has another serious

regret about the paper. “I’d not put XMRV in the title,” Mikovits says.

“We never considered that it would be a single sequence.”

As more negative data poured in, Mikovits and Lombardi became

ever more ardent. Improved techniques now found the virus in

almost every CFS patient, they said.

Patient groups began to see Mikovits as a martyr—a Galileo-like

fi gure fi ghting an all-powerful scientifi c establishment to expose the

truth. But many of her initial supporters had joined the growing camp

of skeptics. “I began comparing Judy Mikovits to Joan of Arc,” Coffi n

says. “The scientists will burn her at the stake, but her faithful follow-

ing will have her canonized.”

Mikovits has her own theory about when Coffi n changed his mind.

She and Lombardi had found evidence, not included in the Science

paper, that XMRV was also linked to autism. On 11 November 2009,

Lombardi presented those data at a meeting at the Cleveland Clinic.

“You don’t talk about autism in the U.S.—it’s too politically charged,”

Mikovits claims Coffi n told him. She believes Coffi n turned against her

that very day. Coffi n confi rms he was upset that Lombardi presented

such preliminary data on such a fraught topic but says, “I did not ‘turn

against’ Judy at that or any other point.”

“Once claims of etiology were made, I just

gasped for breath. My own

experience argued to me

that it’s best to stay away from

this one.”—ROBERT GALLO,

INSTITUTE OF

HUMAN VIROLOGY

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Mikovits further lost credibility with her contribution to an early

study organized by the Blood XMRV Scientifi c Research Working

Group, formed by HHS 1 month after the publication of Lombardi

et al. It included safety experts such as Susan Stramer of the Ameri-

can Red Cross, as well as Mikovits, Alter, Coffi n, and other players

in the XMRV saga. The group had devised a four-stage program that

included sending blinded samples to various labs to see whether they

could detect XMRV. At a meeting on 14 December 2010, the work-

ing group discussed the results of the second stage of its study, which

again did not support Mikovits’s fi ndings. Her lab had found XMRV

in a sample from a healthy person who all labs agreed beforehand was

negative for the virus.

Mikovits had an explanation. The false positive was caused by a

postdoc who mistakenly used the same needle twice to lyse cells and

shear DNA, contaminating the sample. She dismisses the error as triv-

ial, the result of working late at night on a weekend because of repeated

power failures in a new building, coupled with intense pressure from

the working group to get results quickly. “People make mistakes, and

we reported it as a mistake,” she says.

But many researchers threw up their hands: How could WPI botch

a study essential to the survival of its theory? The University of Wash-

ington’s Miller was astonished that Mikovits

didn’t do the critical tests herself and that

they were rushed.

Ruscetti is not bothered that most of

his peers think he’s wrong. “I’ve been there

before,” he says. But he thinks Mikovits has

been treated unfairly. “I’ve been in science for

35 years, and she’s as honest a scientist as I’ve

ever met,” he says.

More setbacks followed. A week after the

working group meeting, Retrovirology pub-

lished four devastating papers online that

together made a compelling case that a contaminant marred both the

Lombardi et al. and Lo-Alter studies. Two reports co-authored by

Coffi n looked for XMRV and MLV in both CFS and prostate cancer

patients; every positive sample they found also harbored mouse DNA,

suggesting that a reagent had become contaminated. Another report

put a fi ner point on it: Researchers discovered MLV—at least 97% sim-

ilar to the Lo-Alter viruses—in a commercially available enzyme used

in a PCR kit. Some speculated that perhaps the patient samples were

handled more than the controls.

The fourth study, led by Greg Towers at University College Lon-

don, was even more damning. It was a follow-up to an earlier report,

published in the April 2009 issue of the Journal of Virology by

Miller’s group, that described the discovery of XMRV in a cell line

called 22Rv1, which was derived from a human prostate cancer patient.

At the time, Miller thought the fi nding supported the theory that the

virus infects humans, as he assumed the original prostate tumor used

to make the cell line harbored XMRV. Now Towers’s team had com-

pared the genetic sequence of XMRV from different 22Rv1 lines with

reported sequences from patients.

The cell line viruses proved ancestral to those in patients. There

was also more genetic diversity in viruses from different 22Rv1 lines

than different patients, precisely the opposite of what should happen if

the virus infected humans: Immune system pressure typically forces a

virus to diversify as a means of escaping attack. Towers concluded that

XMRV never infected humans and that the cell line virus had somehow

contaminated the patient samples.

In March 2011, Coffi n and NCI virologist Vinay Pathak delivered

what many thought was the fi nal blow to XMRV at the Conference on

Retroviruses and Opportunistic Infections in Boston. Working with

Oya Cingöz in Coffi n’s lab, Pathak had analyzed the 22Rv1 cell line

all the way back to its origins, a prostate cancer patient at Case West-

ern Reserve University in Cleveland, Ohio. To create the cell line,

researchers had used a common technique called passaging in which

human tumor cells are grown repeatedly in mice and then harvested.

Early versions of the cell line, still stored in university freezers, had

no XMRV. But two different strains of mice used in the later experi-

ments each harbored DNA that matched half of the virus. This sug-

gested XMRV was created in the 1990s when two viruses combined

in a lab culture, and the widely used cell line spread the virus in many

labs, infecting other cell lines, too. The fi nding furthered doubts that

XMRV had ever infected a human being—let alone had a role in either

CFS or prostate cancer.

The accidental-origin evidence hasn’t convinced Mikovits. “Yes,

that can be an origin of an XMRV,” she says. “But it could have arisen

multiple times. It’s not one sequence.” Pathak contended the chances

that the exact same virus would arise twice were the same as dropping

a quarter from a helicopter fl ying over the Grand Canyon and having it

land on a quarter on the ground.

“It’s all contamination,” Coffi n concluded, which outraged Miko-

vits. “How can John Coffi n shut down research like that?” Mikov-

its shouted during one interview, her blue-gray

eyes shooting fi re. “He’s not God!” She specu-

lates that perhaps the U.S. government, afraid

of the huge consequences of a widespread

XMRV outbreak, was trying to discredit her

work. “We can’t afford another public health

crisis,” she said.

Eventually, two studies that looked for

XMRV in fresh blood samples taken from

patients in the Lombardi paper failed to

fi nd it. Ila Singh, a virologist at the Univer-

sity of Utah in Salt Lake City, consulted with

Mikovits and Lombardi to copy their protocols precisely, and among

100 CFS patients in her study, 14 had tested XMRV-positive by WPI.

Singh and co-workers found 5% of both patients and controls to be

XMRV-positive—a fi nding they traced back to a contaminated enzyme

used in a PCR reaction.

With help from Peterson, UCSF virologist Jay Levy examined 43

patients who tested XMRV-positive at WPI. Peterson says he wanted to

help his former colleagues at the institute. “I went to Jay Levy to prove

them right,” he says. All samples tested negative. Again, Mikovits said

the study was fl awed.

“They didn’t do one thing we did,” Mikovits says.

“We did it exactly the way they did it,” Levy says.

On 31 May, Science published the Levy report and the Coffi n-

Pathak origins study online, along with a so-called Editorial Expres-

sion of Concern by Science Editor-in-Chief Bruce Alberts. “The study

by Lombardi et al. attracted considerable attention, and its publication

in Science has had a far-reaching impact on the community of CFS

patients and beyond,” he wrote, before stating that “the validity of the

study … is now seriously in question.” “What Science giveth, Science

taketh away,” Johnson blogged on Phoenix Rising.

Alberts and Science Executive Editor Monica Bradford had fi rst

suggested that Mikovits and her co-authors retract the paper volun-

tarily. “Science feels it would be in the best interest of the scientifi c

community,” they wrote in a 26 May letter. Mikovits was livid and

questioned Alberts’s motives. “Who wrote that letter? I don’t think it

was Science,” she says. The co-authors thought the retraction request

was premature, too. “What if we walk away from this based on con-

“The virus is real. … I have isolated it from patients. I know it’s there. Believe me.”

—JUDY MIKOVITS,

WHITTEMORE PETERSON INSTITUTE

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tamination and it’s not contamination?” Lombardi asked. “You’ve got to give us time to fi gure this out.”

Alberts stresses that they fl oated the retrac-tion idea because Science already planned to publish the Expression of Concern. “It wasn’t a public call for retraction,” he notes, empha-sizing that the recipients shared it with the media. He also does not think it would have been premature, although he says it’s often a tough call whether to retract a paper. “Ulti-mately, it requires expert judgment and a lot of sensitivity to the issues,” he says. “We had lost confi dence in the results.”

The growing rift between XMRV believers and doubters became painfully obvious at

a June retrovirology meeting in Leuven, Bel-gium, where the two camps literally kept their distance. In the scientifi c sessions, questions were mostly polite and informative. During coffee breaks and a poster session with Bel-gian beer and cheese, Mikovits, her old friend Ruscetti, and De Meirleir, who helped open RedLabs USA, stuck together and barely talked to their scientifi c oppo-nents. The scientifi c debate was grinding to a halt.

“I don’t care if nobody else in the world wants to work on it!” Mikovits exclaimed at one point, rolling her eyes. “Fine, leave us alone!” When Mikovits’s anger subsided, she appeared earnest and even confused by all the criticism. “The virus is real. … I have isolated it from patients. I know it’s there,” she said. “Believe me.”

It wasn’t just that scientists were growing tired of the debate, Wessely says. Some were put off by the “appalling, unforgivable attacks” by some patient advocates on those who criticized WPI’s fi ndings. Wessely says he has received death threats in recent years. “People will rather go over the Niagara in a barrel than ever getting involved in CFS again,” he says.

Silverman notifi ed Science and his collaborators on Lombardi et al. of his discovery that the data he supplied for the paper were wrong. Resequencing samples he tested for the study revealed that somewhere along the line, VP62 had contaminated them. Without casting blame or explaining how the contamination occurred, the partial retraction says two fi gures and a table that reported viral sequences, including one that showed XMRV in the family tree, were “spurious.”

In late August, the Blood Working Group completed its roughly $500,000 study, which conclusively determined that no one need worry about XMRV or MLVs in the blood supply. The nine labs—which included WPI, Ruscetti, and Lo at FDA—each had received blinded samples from 15 negative controls and 15 others who had tested posi-tive for a gammaretrovirus in Lombardi et al. or Lo-Alter. Different teams cultured the virus, looked for antibodies to it, and used PCR to fi sh for DNA. All labs could use whatever assays they chose.

Only WPI and Ruscetti found intermittent evidence of viruses or antibodies in patients, but they also reported similar numbers of pos-itive responses in the negative controls. What’s more, there was no agreement between the two labs on which patient samples tested posi-tive. “What this says, at the very minimum, is that we can’t fi nd it reli-ably in the blood of patients we found it in before,” Ruscetti says.

Ruscetti understands why many of his peers think Lombardi et al. and Lo-Alter have now run their scientifi c course. “It is quite legitimate for those people to say maybe these two papers were wrong,” he says.

But he emphasizes that many scientifi c unknowns remain about XMRV. He points to a study in the Journal of Virology in May that intentionally infected macaques with XMRV. It shows that XMRV moved out of the blood of the monkeys but stayed in tissue reservoirs and that antibod-ies disappeared. “We know nothing about the viral life cycle,” he says.

Blood Working Group member Michael Busch, head of the Blood Systems Research Institute in San Francisco, California, says Ruscetti, Mikovits, Lo, and Alter deserve kudos for participating fully in the study. “I commend them on their scientifi c integrity and commitment to the scientifi c process,” Busch says. “This has been a diffi cult and disappointing process for them and for CFS patients, but hopefully we have all learned lessons that will guide future research and lead to dis-covery of the cause and cure of this disease.”

The results of yet another multilab study are due early next year. Led by Ian Lipkin of Columbia University, the $2.3 million project plans to test 150 samples from CFS patients and create a repository of their blood so that future putative causes of the disease can more easily be tested. Lipkin says the study will continue despite the Blood Working Group’s negative results. “Our study designs are different, our power is different, our subjects are different, and our assays are differ-ent,” Lipkin says. “Whether our results will differ remains to be seen.”

Mikovits and Ruscetti are soldiering on. “As long as there are scientifi c reasons to continue, I think it’s incumbent upon me to do it,” Ruscetti says. “I owe it to both the scientifi c community and the patient community.”

Mikovits says it is irresponsible to dismiss the link between XMRVs and CFS at this point. “Anyone who says this is a lab contaminant has drawn the wrong conclusion and has done a disservice to the public,” she says. Okay, maybe not as many CFS patients have XMRVs as they initially reported, but she’s still convinced that a gammaretrovirus is in at least 20% or 30% of them. “I know of hundreds if not thousands of people with evidence of this infection, from what we’ve done over the last 3 years,” she says. “I don’t know what it means. And I’m gong to keep looking for in vivo reservoirs like the ones seen in the macaques, and I’m going to try to fi gure out mechanisms of pathogenesis, epi-genetics, or other things. I’m not going to stop studying it.”

–JON COHEN AND MARTIN ENSERINK

“People will

rather go over

the Niagara in a

barrel than ever

getting involved in

CFS again.”—SIMON WESSELY,

KING’S COLLEGE

LONDON

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