16S nanopore sequencing accurately detects intraamniotic infection

16S nanopore sequencing accurately detects intraamniotic infection | Image Credit: © Rattanachat - © Rattanachat - stock.adobe.com.

The 16S nanopore sequencing is effective for predicting intraamniotic infection, a key factor of preterm birth, according to a recent study published in the American Journal of Obstetrics & Gynecology.¹

Preterm birth is the main source of perinatal morbidity and mortality globally, with over 2 in 3 preterm deliveries occurring after spontaneous labor onset.² In cases of intraamniotic infection, microorganisms reach the amniotic cavity, leading to a local inflammatory response.¹

“Diagnosis of intraamniotic infection is important in order to optimize the treatment of mothers and newborns, given recent evidence that intraamniotic infection and intraamniotic inflammation can be successfully treated,” wrote investigators.

Assessing intraamniotic infection

The prospective study was conducted to evaluate the diagnostic performance of the 16S ribosomal deoxyribonucleic acid (rDNA) nanopore sequencing method for intraamniotic infection. This method identified bacteria by confirming the presence of the 16S rDNA gene, which can be used to confirm bacteria across taxonomic levels.

Participants included adult women with a singleton pregnancy diagnosed with spontaneous labor, either with intact membranes or preterm prelabor rupture of the membranes (PPROM). Exclusion criteria included refusing amniocentesis, underlying infectious diseases, intrauterine fetal death, and emergent obstetrical conditions.

Patients receiving genetic amniocentesis in the mid-trimester with a negative amniotic fluid culture and 16S rDNA sequencing result were classified as controls. Amniotic fluid was obtained through transabdominal ultrasound-guided amniocentesis.

Criteria for infection and inflammation

A positive culture for bacteria or the presence of microbial footprints for bacteria during 16A rDNA sequencing indicated microbial invasion of the amniotic cavity (MIAC). Amniotic fluid interleukin 6 concentration of 2.6 ng/mL or greater indicated intraamniotic infection.

Four subgroups of amniotic fluid culture were developed. These included no infection or inflation, MIAC without intraamniotic inflation, sterile intraamniotic inflammation, and intraamniotic infection.

A triple antibiotic regimen was provided to patients with intraamniotic inflammation under 32- to 34-weeks’ gestation, with delivery conducted in cases of clinical chorioamnionitis. In those at least 32- to 34-weeks’ gestation, delivery was considered for intraamniotic infection and intraamniotic inflammation.

Infection and inflammation findings

There were 52 participants included in the final analysis, 55.8% of whom had intact membranes and 44.2% had PPROM. A median gestational age of 30.9 weeks was reported at amniocentesis vs 33 weeks at delivery. Acute inflammatory lesions in the placenta were reported in 33.3% of patients, while 51.9% had no intraamniotic infection or inflammation.

Sterile intraamniotic inflammation was identified in 23.1% of patients, intraamniotic infection in 17.3%, and MIAC in 7.7%. The greatest number of bacterial reads was reported in patients with intraamniotic infection.

In cases of intraamniotic infection, Ureaplasma spp was found in 33%, making it the most common microorganisms identified through culture or 16S sequencing. This was followed by Gardnerella vaginalis, Klebsiella pneumoniae, Pseudomonas veronii, Streptococcus anginosus, Streptococcus mitis, and Actinotignum schaalii all in 11%.

While polymicrobial infection was not identified through cultivation, 16S rDNA sequencing successfully identified polymicrobial infection in 33% of patients with intraamniotic infection. Additionally, unlike Sanger sequencing, nanopore sequencing successfully identified specific microorganisms at the species level.

Implications

In patients with sterile intraamniotic infection, negligible read counts below 100 or the presence of sequences linked to known background bacterial species were identified through 16S nanopore sequencing. Negative amniotic fluid culture was identified in those with mid-trimester amniocentesis through 16S Sanger sequencing.

Overall, the results highlighted high sensitivity, specificity, accuracy, negative predictive value, and a positive likelihood for identifying intraamniotic infection in patients with preterm delivery.

“We foresee that the combination of 16S nanopore sequencing and rapid assessment of intraamniotic inflammation… is a step toward the accurate identification of true intraamniotic infection,” wrote investigators.

References

1. Chaemsaithong P, Romero R, Pongchaikul P, et al. The rapid diagnosis of intraamniotic infection with nanopore sequencing. Am J Obstet Gynecol. 2025;233:193.e1-18. doi:10.1016/j.ajog.2025.02.011
2. Goldenberg RL, Culhane JF, Iams JD, Romero R. Epidemiology and causes of preterm birth. Lancet. 2008;371(9606):75-84. doi:10.1016/S0140-6736(08)60074-4