Infections Linked to Premature Births More Common Than Thought, Stanford Study Finds

STANFORD, Calif.--(Business Wire)--
Previously unrecognized and unidentified infections of amniotic
fluid may be a significant cause of premature birth, according to
researchers at the Stanford University School of Medicine.

An analysis of amniotic fluid from women in preterm labor
indicated that 15 percent of the fluid samples harbored bacteria or
fungi -- an increase of 50 percent over previous estimates. The
heavier the burden of infection, the more likely the women were to
deliver younger, sicker infants.

"If we could prevent these infections in the first place, or
detect them sooner, we might one day be able to prevent some of these
premature births," said research associate Dan DiGiulio, MD, who
conducted the study in the laboratory of senior author David Relman,
MD. About 12 percent of all births in this country are premature and
the frequency of premature birth is increasing.

"To find that this amniotic compartment, which we have
traditionally viewed as somewhat sacrosanct, is infected significantly
more often than we thought is a little shocking," said Relman,
professor of infectious disease and of microbiology and immunology.
Previous analyses of the same samples using a different, more
conventional method had concluded that only about 10 percent were
infected.

DiGiulio and Relman collaborated with researchers at Wayne State
University in Detroit, the Detroit Medical Center and the Eunice
Kennedy Shriver National Institute of Child Health and Human
Development to conduct the research. DiGiulio is the first author of
the study, which will be published in the Aug. 26 issue of PLoS-ONE.

Relman received a 2006 Pioneer Award from the National Institutes
of Health and is using the proceeds to characterize the microbial
communities indigenous to humans and understand the roles of these
communities in health and disease.

For this study, DiGiulio used a highly sensitive technique called
polymerase chain reaction, or PCR, to track down micro-organisms in
amniotic fluid samples collected from 166 women in preterm labor at
the Detroit Medical Center between October 1998 and December 2002. Of
these, 113 women went on to deliver their infants prematurely.
Although not all these women were infected, such microbial invasion is
thought to contribute significantly to the overall number of preterm
births.

Previous efforts to conduct a census of bacteria, fungi and other
living organisms in the fluid relied primarily on culturing, or
growing, the invaders in the laboratory after withdrawing some of the
liquid through a needle inserted into the amniotic cavity. But not
every microbe is amenable to such a rough-and-tumble change of
quarters.

"We've become very good at growing some bacterial species -- what
might be called the 'weeds' of the microbial world," said DiGiulio.
"But many other species can't be cultured by current methods." In
contrast, the PCR technique used by the researchers tracks down and
copies small portions of DNA encoding a cellular component called
ribosomal RNA that is shared by all living creatures -- kind of like
screening for "all people with fingers." Small differences among the
fingerprints identify individual species.

In this way, the researchers cast a wide net and got the results
to show for it. They used both PCR and laboratory cultures to
determine that 25 of the 166 samples were infected with either
bacteria or fungi. Seventeen bacterial and one fungal species were
identified in the positive samples -- far more than the 11 species
found by conventional analysis -- including some not previously
suspected to play a role in preterm delivery. Furthermore, the
investigation turned up a novel micro-organism that may represent a
previously unknown species.

One of the study's collaborators was Roberto Romero, MD, chief of
the Perinatology Research Branch of the Eunice Kennedy Shriver
National Institute of Child Health and Human Disease. Romero is also a
professor of molecular obstetrics and genetics at Wayne State
University. He is well-known for his research showing that infection
is a common and important cause of preterm labor and delivery, and
that affected fetuses may be predisposed to short- and long-term
complications.

Infection of the amniotic fluid likely contributes to preterm
delivery by causing an inflammatory response in the pregnant woman. It
occurs most commonly either by microbes from the vagina that
infiltrate the amniotic sac, or by microbes that travel through the
mother's bloodstream from other parts of her body, such as the mouth.
In fact, both gum disease and bacterial vaginosis have been shown to
increase a woman's risk of delivering her baby prematurely.

DiGiulio and Relman found that all the laboring women whose
samples were positive by either PCR or by culture delivered their
infants prematurely. In addition, all those who were positive by both
methods delivered their babies within one day of the initial fluid
collection, and 68 percent of those that were positive by only one,
but not the other, method delivered within the same time period.

They also found that 27 percent of samples from infected women who
delivered their infants before 25 weeks gestation were positive only
by PCR; cultures from these samples falsely indicated they were
uninfected. Because only about 30 percent of infants in this age group
survive their early birth, it is extremely important to be able to
accurately diagnose infection in these women.

Finally, as the burden of infection increased, the average
gestational age of the newborns decreased. That is, a woman with many
copies of bacterial or fungal rRNA, which suggests the presence of
many bacterial or fungal cells, was likely to deliver a very young,
very sick infant. Relman and DiGiulio are now collaborating with
Romero's group on a much larger study of fluid collected during
routine amniocenteses at about the 20th week of pregnancy. They hope
to determine whether infections can be detected before the onset of
preterm labor, which could lead to new prevention or treatment
strategies.

"Although we know now that humans should respect and even cherish
their microbial symbionts," said Relman, "this type of infection
reflects a situation in which our microbial companions have
overstepped their bounds. Something has gone awry with this balance."

DiGiulio worked with Elisabeth Bik, PhD, a research associate in
Relman's group at Stanford and the Veterans Affairs Palo Alto Health
Care System, as well as colleagues who work with Romero at Wayne State
University School of Medicine and the National Institute of Child
Health and Human Development.

In addition to Relman's NIH award, this work was supported by
NICHD.

The paper is available online at
http://dx.plos.org/10.1371/journal.pone.0003056.

Stanford University Medical Center integrates research, medical
education and patient care at its three institutions -- Stanford
University School of Medicine, Stanford Hospital & Clinics and Lucile
Packard Children's Hospital at Stanford. For more information, please
visit the Web site of the medical center's Office of Communication &
Public Affairs at http://mednews.stanford.edu.

Stanford University Medical Center
Krista Conger, 650-725-5371 (Print Media)
kristac@stanford.edu
Erin Digitale, 650-724-9175 (Print Media)
digitale@stanford.edu
Margarita Gallardo, 650-723-7897 (Broadcast Media)
mjgallardo@stanford.edu

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