Intra-Amniotic Infection/Inflammation and the Fetal Inflammatory Response Syndrome

Introduction

Infection is the greatest killer in human history. The microbial theory of disease is probably the greatest advance in medicine, for it has made possible the identification of the cause of many diseases, the development of diagnostic modalities (i.e., cultivation of microorganisms), treatment with antimicrobial agents, and the development of vaccines. The achievements of the microbial theory of disease in medicine and public health are unparalleled with regard to any other contribution in medicine. The formulation of the Koch postulates to establish causality between a microbe and a disease provided a major breakthrough that has influenced the philosophy of science. Although during the 20th century some considered that infectious diseases had been conquered, the pandemic of coronavirus disease 2019 (COVID-19), epidemics of acquired immunodeficiency syndrome (AIDS), Ebola virus, swine flu, and the like tell a different story. The recent understanding that infections during pregnancy may predispose to schizophrenia, autism spectrum disorders, and cerebral palsy ^, in the offspring suggests that insults occurring during intrauterine life (infection-induced inflammation) can have lasting effects.

Fetal and neonatal physicians and scientists have been interested in intra-amniotic infection as a cause for preterm labor and delivery, neonatal morbidity and mortality, and long-term disorders. This chapter will focus largely on the microbiology, the innate immune response, and the fetal inflammatory response syndrome (FIRS).

Birth: Transition from a Sterile to a Nonsterile Environment

Four major transitions occur at birth: (1) emergence from an aquatic environment where oxygen is acquired through the placenta to a dry environment in which respiratory exchange occurs through the lungs; (2) adaptation from a warm environment in which the fetus has a temperature that is 1°C higher on average than that of the mother to a cooler environment at room temperature; (3) exchange of a continuous supply of nutrients through the placenta for intermittent feedings during the neonatal period; and (4) transition from a sterile bacterial environment to the establishment of the neonatal microbiome (e.g., gut, respiratory tract, skin).

The fetus normally lives in an environment devoid of bacteria, as determined by cultivation and molecular microbiologic techniques of amniotic fluid. ^, Therefore, birth represents a critical stage for acquisition of the first microbiota. During labor, the fetus becomes exposed to the vaginal microbiota and such bacteria become the pioneer microorganisms that invade the formerly sterile body of the infant to establish the first neonatal microbiota. ^, Such microbiota may differ when an infant is born via vaginal or cesarean delivery.

Under pathologic conditions, however, microbial invasion of the amniotic cavity due to bacteria, fungi, and viruses can predispose to fetal infection. Microorganisms can enter the fetus through different sites, including the mucous membranes of the airways, gastrointestinal tract, tympanic membrane, the conjunctiva, or skin. Once bacteria are in contact with the mucous membranes, they can gain access to the fetal circulation and cause bacteremia, which then can lead to sepsis, septic shock, or even death. The fetal immune system, after the second trimester of pregnancy, is capable of mounting both an innate and an adaptive immune response, which play a key role in host defense against infection. Indeed, recent studies show that the amniotic fluid harbors innate and adaptive immune cells of fetal origin, which play a central role in host defense against microbes invading the amniotic cavity.

Microbial invasion of the amniotic cavity has been causally linked to preterm labor with intact or ruptured membranes. One-third of preterm neonates are born to mothers with microbial invasion of the amniotic cavity. ^{, ,} Moreover, the earlier the gestational age at delivery, the greater the frequency of intra-amniotic infection and the greater the burden of disease. ^,

Nomenclature: Microbial Invasion of the Amniotic Cavity, Intra-Amniotic Inflammation, Intra-Amniotic Infection, Clinical Chorioamnionitis, Acute Histologic Chorioamnionitis, and Funisitis

Table 12.1 and Fig. 12.1 define terms often used to refer to entities related to intra-amniotic infection. Microbial invasion of the amniotic cavity is defined as the presence of organisms in the samples of amniotic fluid retrieved by transabdominal amniocentesis and detected by cultivation methods and/or molecular microbiologic techniques. Intra-amniotic inflammation is the presence of an inflammatory response in the amniotic cavity, which can be detected by the presence of inflammatory cells, for instance, white blood cells, in an elevated concentration of an inflammatory mediator, such as interleukin (IL)-6, ³⁶ or in an elevated concentration of matrix metalloproteinase (MMP). Rapid tests that can determine these biomarkers have become available recently as point-of-care tests, and results can be obtained in less than 20 minutes without specialized laboratory equipment.

The term intra-amniotic infection is used when there is a combination of microbial invasion of the amniotic cavity and intra-amniotic inflammation. Most cases of microbial invasion of the amniotic cavity without evidence of inflammation are due to specimen contamination by skin flora, laboratory bacteria, or reagents. When intra-amniotic inflammation is present in the absence of microorganisms, the condition is referred to as sterile intra-amniotic inflammation. ^, This term should be reserved for samples that are negative for microorganisms detected by both cultivation methods and molecular microbiologic techniques. ^, Sterile intra-amniotic inflammation has been attributed to “danger signals” (or alarmins ) that are released during cellular stress or cell necrosis.

Clinical chorioamnionitis refers to a syndrome characterized by the presence of a maternal fever (temperature ≥37.8°C or ≥38.0°C) and two or more of the five following clinical signs: (1) maternal tachycardia (heart rate >100 beats/min); (2) fetal tachycardia (heart rate >160 beats/min); (3) uterine tenderness; (4) purulent or malodorous amniotic fluid or vaginal discharge; and (5) maternal leukocytosis (white blood cell count >15,000/mm ³ ).

Clinical chorioamnionitis represents the clinical manifestation of a maternal systemic inflammatory response and is present in only 10% to 20% of all patients with proven intra-amniotic infection. Acute histologic chorioamnionitis is the presence of neutrophils in the chorioamniotic membranes or the chorionic plate and represents a maternal host response. Funisitis is inflammation of the umbilical cord (umbilical vein, artery, and Wharton’s jelly) and is a fetal host response. ^, Chorionic vasculitis consists of inflammation of the fetal vessels on the surface of the chorionic plate and is evidence of fetal inflammation. Acute histologic chorioamnionitis, more common than clinical chorioamnionitis, is the pathologic expression of intra-amniotic inflammation. ^{, ,} However, acute histologic chorioamnionitis should not be considered synonymous with intra-amniotic infection, since a fraction of patients with sterile intra-amniotic inflammation have acute histologic chorioamnionitis and, sometimes, funisitis. The term acute refers to a specific inflammatory lesion in which the predominant cell is the neutrophil. Chronic inflammatory lesions of the placenta are characterized by the infiltration of lymphocytes, plasma cells, and macrophages. Although chronic inflammatory lesions could be due to specific infectious agents (such as viruses), the main cause of chronic inflammatory lesions is maternal antifetal rejection, and interested readers are referred to a recent review by the authors on this subject.

Pathways for Intra-Amniotic Infection

Microorganisms may gain access to the amniotic cavity through three main pathways : (1) ascension from the vagina and cervix ^, ; (2) hematogenous dissemination through the placenta (transplacental infection) ^, ; and (3) accidental introduction at the time of invasive procedures, such as amniocentesis, percutaneous fetal blood sampling, chorionic villus sampling, or shunting.

The most common pathway of intra-amniotic infection is the ascending route, ^{, , ,} evidenced as follows:

• 1.

  The bacteria isolated in the amniotic fluid of patients with intra-amniotic infection are frequently found in the vagina. ^,

• 2.

  In twin gestations, acute histologic chorioamnionitis, more common in the first-born twin, is rarely seen in the second twin. Moreover, when intra-amniotic infection is detected, the presenting sac is nearly always involved.

• 3.

  In virtually all cases of congenital pneumonia, inflammation of the chorioamniotic membranes is present. ^{, ,}

Stages of Ascending Infection

The stages of ascending infection are listed in Fig. 12.2 . Stage 1 corresponds to a change in the vaginal/cervical microbial flora or to the presence of pathologic organisms (e.g., Neisseria gonorrhoeae ) in the cervix. Some forms of bacterial vaginosis may be an early manifestation of this initial stage. An abnormal vaginal microbiota (even in the absence of bacterial vaginosis) can correspond to the first stage and predispose to preterm delivery. The specific changes in the vaginal microbiota have been recently described by using sequence-based techniques. ^, In stage 2 , microorganisms gain access to the uterine cavity and reside in the lower pole of the uterus between the membranes and chorion, where they can elicit a localized inflammatory reaction. In stage 3, bacteria have penetrated the chorion and the amnion in the lower pole of the uterus into the amniotic cavity, leading to microbial invasion of the amniotic cavity and intra-amniotic infection when an inflammatory response is established in the amniotic fluid. Rupture of the membranes is not a prerequisite for intra-amniotic infection because microorganisms are capable of crossing intact membranes. Finally, in stage 4 , bacteria may gain access to the fetus through different sites. Aspiration of infected fluid by the fetus may lead to congenital pneumonia. Otitis, conjunctivitis, and omphalitis may occur by the direct spreading of microorganisms from infected amniotic fluid. Seeding from any of these sites to the fetal circulation may result in fetal bacteremia and sepsis.

The traditional concept has been that bacteria invade the decidua from the lower genital tract and, from there, are localized to the space between the chorion and amnion before invading the amniotic cavity. ^, However, molecular data bring into question whether most cases of intra-amniotic infection are preceded by a stage in which the infection is in the decidua or in the chorioamniotic space.

Microbiology of Intra-Amniotic Infection

The most common microbial isolates from the amniotic cavity of women with intra-amniotic infection are Ureaplasma spp. ( Ureaplasma parvum and Ureaplasma urealyticum ), Fusobacterium spp., and Mycoplasma hominis. ^a

a References 29, 31, 53, 102, 121, 128–136.

These microorganisms are frequently found as members of the vaginal microbiome in normal pregnant women. Why these specific microorganisms ascend into the amniotic cavity and not others, which are more frequently present, such as Lactobacilli spp., is unknown. Table 12.2 displays the most frequent microorganisms found in the amniotic fluid of patients with intra-amniotic infection. Most microorganisms are also found in the lower genital tract of normal pregnant women.

In approximately 50% of patients with intra-amniotic infection, more than one microorganism are isolated from amniotic fluid. The inoculum size differs considerably, and in 71% of cases, more than 10 ⁵ colony-forming units per milliliter are found. Chlamydia trachomatis is rarely found in the amniotic fluid.

Viral invasion of the amniotic cavity has been found in 2.2% of women (16 of 729) undergoing second-trimester amniocentesis for genetic indications. Human herpes virus 6 is the most common organism, followed by human cytomegalovirus, parvovirus B19, and Epstein-Barr virus. Studies of the presence of viral genomes in amniotic fluid that used targeted polymerase chain reaction (PCR) approaches showed that viruses are extremely rare in patients with preterm prelabor rupture of the membranes (PROM). ^, Characterization of the amniotic fluid virome is a frontier.

Frequency of Microbial Invasion of the Amniotic Cavity Due to Bacteria