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Microbial burden and inflammasome activation in amniotic fluid of patients with preterm prelabor rupture of membranes - PubMed

  • ️Wed Jan 01 2020

Microbial burden and inflammasome activation in amniotic fluid of patients with preterm prelabor rupture of membranes

Kevin R Theis et al. J Perinat Med. 2020.

Abstract

Background Intra-amniotic inflammation, which is associated with adverse pregnancy outcomes, can occur in the presence or absence of detectable microorganisms, and involves activation of the inflammasome. Intra-amniotic inflammasome activation has been reported in clinical chorioamnionitis at term and preterm labor with intact membranes, but it has not yet been investigated in women with preterm prelabor rupture of membranes (preterm PROM) in the presence/absence of detectable microorganisms. The aim of this study was to determine whether, among women with preterm PROM, there is an association between detectable microorganisms in amniotic fluid and intra-amniotic inflammation, and whether intra-amniotic inflammasome activation correlates with microbial burden. Methods Amniotic fluids from 59 cases of preterm PROM were examined for the presence/absence of microorganisms through culture and 16S ribosomal RNA (rRNA) gene quantitative real-time polymerase chain reaction (qPCR), and concentrations of interleukin-6 (IL-6) and ASC [apoptosis-associated spec-like protein containing a caspase recruitment domain (CARD)], an indicator of inflammasome activation, were determined. Results qPCR identified more microbe-positive amniotic fluids than culture. Greater than 50% of patients with a negative culture and high IL-6 concentration in amniotic fluid yielded a positive qPCR signal. ASC concentrations were greatest in patients with high qPCR signals and elevated IL-6 concentrations in amniotic fluid (i.e. intra-amniotic infection). ASC concentrations tended to increase in patients without detectable microorganisms but yet with elevated IL-6 concentrations (i.e. sterile intra-amniotic inflammation) compared to those without intra-amniotic inflammation. Conclusion qPCR is a valuable complement to microbiological culture for the detection of microorganisms in the amniotic cavity in women with preterm PROM, and microbial burden is associated with the severity of intra-amniotic inflammatory response, including inflammasome activation.

Keywords: PPROM (preterm prelabor rupture of membranes); culture; microbial invasion of the amniotic cavity; quantitative real-time PCR (qPCR); sterile intra-amniotic inflammation.

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Conflict of interest statement

Conflicts of interest: The authors declare that there are no conflicts of interest.

Figures

Figure 1.
Figure 1.. Association between microbiological culture and 16s rRNA gene qPCR in amniotic fluid.

Cycle of quantification (Cq) of background technical controls and amniotic fluid samples from women with preterm PROM based on the presence/absence of positive microbial culture. Median values are indicated. Statistical results are from Mann-Whitney U tests. Dotted squares represent the absence of microbial detection by 16S rRNA gene qPCR in culture positive/negative amniotic fluid samples. N = 20–37 per group.

Figure 2.
Figure 2.. Microbial burden and intra-amniotic inflammation.

Cycle of quantification (Cq) of background technical controls and amniotic fluid samples based on the presence/absence of positive microbiological culture and intra-amniotic inflammation (IL-6 concentrations > 2.6 ng/mL). Median values are indicated. Statistical results are from Mann-Whitney U and Kruskal-Wallis tests. Sequential Bonferroni corrections were applied to all post hoc pairwise comparisons. The dashed line indicates the lowest Cq value of any background technical control, which was used to define the cutoff for a positive 16S signal (Cq value < 34.66 cycles). N = 12–19 per group.

Figure 3.
Figure 3.. Correlation between microbial burden and extracellular ASC in amniotic fluid.

Extracellular ASC concentration in relation to the cycle of quantification (Cq) of amniotic fluid samples. Categorization of amniotic fluid samples is based on the presence/absence of positive microbiological culture and/or positive bacterial 16S rRNA gene qPCR, and intra-amniotic inflammation (IL-6 concentrations > 2.6 ng/mL). The statistical result is from a Spearman’s rank-order correlation test. The regression line is indicated. N = 8–19 per group.

Figure 4.
Figure 4.. Extracellular ASC concentrations in amniotic fluid of women with preterm PROM in the presence/absence of positive microbial detection and intra-amniotic inflammation.

Extracellular ASC concentrations of amniotic fluid samples based on the presence/absence of positive microbial culture and/or positive bacterial 16S rRNA gene qPCR, and intra-amniotic inflammation (IL-6 concentrations > 2.6 ng/mL). Median values are indicated. Statistical results are from Kruskal-Wallis and Mann-Whitney U tests. Amniotic fluid categories marked by different letters were statistically different after sequential Bonferroni corrections were applied. N = 8–19 per group.

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