The rate of organism recovery in this study is so high that we wondered how much reflects colonization and how much can be attributed to contamination. We offer findings from this and other studies to support the view that many of the microorganisms isolated during this study represent colonization of the placenta parenchyma.
The methods used during this study were designed to recover both obligately anaerobic and facultative bacterial species. Previous studies have shown that both obligate anaerobes and facultative species are readily recovered from frozen tissues and that anaerobes that are part of normal vaginal microflora can survive 4 or more hours of exposure to room air. 19-22
Since tissues removed from a systemic supply of oxygen have a much lower oxygen tension than room air (20% O2
), the tissue itself serves as a protective barrier for presumed oxygen sensitive bacteria. We believe that the methods employed were adequate for recovering microorganisms when present. Moreover, the use of PCR on a subset of specimens with both universal and specific primers did not identify microbial DNA in culture-negative or culture-positive specimens.13
Placentas from pregnancies that ended with preterm labor had recovery rates that declined with increasing gestational age. If the presence of microorganisms within the placenta were simply a reflection of contamination, the rate of recovery should be the same regardless of gestational age. Because the duration of labor did not vary appreciably with gestational age, the observed gestational age-organism recovery relationships are unlikely to be confounded by duration of labor. In this study, the chorion of placentas from pregnancies that ended with preterm labor had much higher rates of microorganism recovery than placentas from pregnancies that ended early because of increasingly severe preeclampsia. The data suggest that recovery of microorganisms from placental tissue convey biologically important information. Indeed, our findings are consistent with other evidence that infectious/inflammatory phenomena contribute to the onset of labor before the third trimester. 23, 24
Placentas that had neutrophils in fetal stem vessels within the chorion or in the vessels of the cord (identified as fetal vasculitis) had higher rates of microorganism recovery than placentas without fetal vasculitis. These histologic characteristics are considered manifestations of a fetal inflammatory response, a recognized antecedent of preterm labor. 24
The numbers of colony forming units/gram of tissue also exceed the levels generally associated with contamination. Contamination of tissue specimens obtained after antiseptic preparation usually occurs with recovery of very low levels of microorganisms (e.g. fewer than 102 colony forming units (cfu)/gram of tissue). The uniformly high median counts of organisms we recovered suggest that most of the microorganisms isolated were not contaminants.
The rate of microorganism isolation was higher in placentas delivered vaginally than in those delivered by CS. Because some of the difference in isolation rate might be due to contamination as the placenta passed through the vagina, we restricted our attention to placentas delivered by CS. Nevertheless, some of our inferences might also apply to vaginally delivered placentas.
The sterile technique to obtain sterile tissue included lifting and cutting open the amnion with sterile instruments, and peeling away this protective cover to expose the chorion. A second set of sterile instruments was then used to sample the chorion and underlying trophoblast. These efforts should have kept contamination to a minimum. No single species or small group of species consistent with the distribution of species found on skin or on the vaginal epithelium accounted for a sizable proportion of microorganisms isolated from placentas following preterm labor, or from placentas characterized by low gestational age or fetal vasculitis. This is what would be expected with low-virulence organisms, and what has been seen with cultures of amniotic fluid and membranes from preterm pregnancies. 23, 24
In a separate study of term placentas, we used the same sample collection and culture methods for CS delivered placentas and found that only 17% (2/12) of specimens yielded a microorganism when cultured. This is similar to our recovering organisms from approximately 20% of preeclamptic placentas delivered before the 28th post-menstrual week. We also found in our small study of term placentas that organisms were not recovered in the absence of labor (unpublished data).
Because bacteria-initiated inflammation has not been associated with severe preeclampsia, CS delivered placentas of preeclampsia pregnancies are considered least likely to yield a microorganism. Yet, a microorganism was recovered from almost a quarter of these placentas. In 83% of these cases, the isolate was a single microorganism, such as Propionibacterium, E. coli, coagulase negative staphylococci, Group D streptococci, and alpha hemolytic streptococci. While Propionebacterium sp., and coagulase negative staphylococci may be considered as possible contaminants from skin, E. coli, Group D streptococci and alpha hemolytic streptococci are not commonly members of the cutaneous microflora, nor would any of these species be expected to survive the routine antiseptic preparation before a Caesarian delivery. Consequently, we cannot estimate accurately how much contamination as opposed to true colonization occurs in CS-delivered placentas.
We acknowledge that it is provocative to propose that the presence of microorganisms in these specimens represents colonization and that those organisms in the immediate environment are most likely to colonize the placenta. On the other hand, antibiotics can promote endometrial resolution, but not the eradication, of gram-variable rods such as G. vaginalis
, as well as new bacterial acquisitions. 25
More than 80% of 820 endometrium cultures obtained three months after delivery yielded a microorganism. 5
The authors of that study argued that contamination could not account for all that they saw because half of these specimens had plasma cell endometritis, and because the microorganisms recovered did not match the distribution of species in the vagina, the most probable source of the putative contamination. In light of our study, and the others that have recovered organisms from the endometrium 1-5, 6-7
, we feel the time has come to recognize that the uterus is not a sterile organ.
Until recently, it was thought that microorganisms from the vagina passed the barrier created by the fetal membranes thereby gaining access to the amniotic sac, 26-28
and that maternal immunosuppression that allowed a tolerance of the fetal allograft also prevented an intense immunologic response to these bacteria. 29
What are these microorganisms doing in the amniotic sac and placenta? The deleterious effects of colonization appear to provoke an inflammatory response that results in preterm labor and delivery. Such an undesirable result might occur if the placenta fails at its filtering/removal function and lets microorganisms survive and invade feta tissues. 30
Indeed, one could argue that the higher levels of bacterial isolation from vaginally delivered placentas simply indicate that the fetus senses a hostile microbiologic environment and signals the host in such a way as to escape from this environment. 31
Because the normal route of delivery is through the birth canal, the delivered fetus requires some rudimentary protection in place as it passes through an area containing large numbers of microorganisms. 32
Bacterial colonization of the placenta might promote normal development of the fetal immune system without harm to fetus or mother. For example, Lactobacillus
sp., and coagulase negative Staphylococcus
sp. are among the least pathogenic, most commonly isolated and most numerous of vaginal microflora microorganisms. Both groups of microorganisms have cell wall constituents, such as peptidoglycan and lipoteichoic acid that are considered highly immunogenic.33-35
Small amounts of these cell wall constituents have the potential to promote maturation of the fetal immune system during pregnancy. Since much of the developing neonatal immune system is provoked by GI microflora following birth,36
might placenta bacteria play a role in immune development before birth?
Placenta bacteria might also be beneficial in other ways. One group of commentators went so far as to suggest, “The emerging picture is that microbial-host interactions in the endometrial cavity are important for reproductive success”.37
In addition to those microorganisms considered to be non-pathogens, other species including Prevotella
sp., Gardnerella vaginalis, Ureaplasma urealyticum
sp., may be both important immunomodulators, and capable of escaping the placental/maternal defense mechanisms to cause an outright inflammatory process.
The axiom that single microorganism cultures are more likely to be invasive pathogens is not born out by the findings of . Fifty-six percent of all cultures that yielded coagulase negative staphylococci from labor-initiated deliveries were single-microorganism cultures (). This is the exact same percent of all E. coli
cultures that were unaccompanied by another microorganism. Polymicrobial infections do occur, particularly adjacent to surfaces with complex microflora 38
and often include commensal microbial species. 39,40
Perhaps the most attractive feature of this study is the large number of placentas cultured. This has allowed stratification of the data by potential confounding variables. An added attraction, and one that makes this study unique, is the use of placenta parenchyma for culture and not the more commonly cultured amniotic and chorionic membranes or amniotic fluid. If one assumes that placenta parenchyma is sterile, then finding microorganisms within the tissue specimens is an important observation.
Previous reports of the association of bacteria with preterm labor have relied on the recovery of microorganisms from amniotic fluid or the membranes.17, 41-44
To our knowledge, this is the first report of an analysis of bacteria within placenta parenchyma at the time of preterm delivery.
In summary, the increased rates of microorganism recovery from placenta parenchyma associated with low gestational age, preterm labor and fetal vasculitis suggest that an appreciable proportion of the microorganisms recovered contribute to preterm labor and the fetal inflammatory response, which might be an intermediary between the microorganism and preterm labor. In addition, the finding of presumed non-pathogenic microorganisms within the placenta parenchyma is an important new observation.