title: Uncultivated bacteria as etiologic agents of intra-amniotic inflammation leading to preterm birth.
abstract: Intra-amniotic infection and inflammation are major causes of preterm birth (PTB). However, intra-amniotic inflammation is often detected in the absence of infection. This may partly be due to the culturing methods employed in hospital laboratories, which are unable to detect the uncultivated species. In this study, intra-amniotic microbial infections associated with PTB were examined by both culture and 16S rRNA-based culture-independent methods and were corroborated by the presence of intra-amniotic inflammation. Amniotic fluid (AF) specimens from 46 pregnancies complicated by PTB and 16 asymptomatic women were analyzed. No bacterial DNA was amplified in AF collected from the asymptomatic women. Among the 46 samples associated with PTB, bacterial DNA was amplified from all (16/16) of the culture-positive samples and 17% (5/30) of the culture-negative samples. In the culture-positive group, additional species were detected in more than half (9/16) of the cases by PCR and clone analysis. Altogether, approximately two- thirds of the species detected by the culture-independent methods were not isolated by culture. They included both uncultivated and difficult-to-cultivate species, such as Fusobacterium nucleatum, Leptotrichia (Sneathia) spp., a Bergeyella sp., a Peptostreptococcus sp., Bacteroides spp., and a species of the order Clostridiales. To examine intra-amniotic inflammation, an AF proteomic fingerprint (mass-restricted score) was determined by surface-enhanced laser desorption ionization-time-of-flight mass spectrometry. Inflammation was detected in all five samples which were culture negative but PCR positive. Women who were PCR positive more often had elevated interleukin-6 levels in their AF, histological chorioamnionitis, and funisitis and delivered neonates with early-onset neonatal sepsis. Previously unrecognized, uncultivated, or difficult-to-cultivate species may play a key role in the initiation of PTB.
authors.affiliation: Department of Periodontics, School of Dental Medicine, Case Western Reserve University, 10900 Euclid Avenue, Cleveland, OH 44106-4905, USA. yiping.han@case.edu
authors.forename: Yiping W
authors.initials: YW
authors.lastname: Han
authors.forename: Tao
authors.initials: T
authors.lastname: Shen
authors.forename: Peter
authors.initials: P
authors.lastname: Chung
authors.forename: Irina A
authors.initials: IA
authors.lastname: Buhimschi
authors.forename: Catalin S
authors.initials: CS
authors.lastname: Buhimschi
mesh_terms: D000328:Adult; D000650:Amnion / Q000382:microbiology*; D000653:Amniotic Fluid / Q000737:chemistry; D001419:Bacteria / Q000145:classification* / Q000235:genetics / Q000254:growth & development / Q000302:isolation & purification*; D001424:Bacterial Infections / Q000150:complications*; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000235:genetics; D004275:DNA, Ribosomal / Q000235:genetics; D005260:Female; D006801:Humans; D007231:Infant, Newborn; D013058:Mass Spectrometry; D016133:Polymerase Chain Reaction; D011247:Pregnancy; D011251:Pregnancy Complications, Infectious; D047928:Premature Birth / Q000209:etiology*; D020543:Proteome / Q000032:analysis; D012336:RNA, Ribosomal, 16S / Q000235:genetics
chemical_list: D004269:DNA, Bacterial; D004275:DNA, Ribosomal; D020543:Proteome; D012336:RNA, Ribosomal, 16S