Fetal and perinatal infections: A consecutive study

Path. Res. Pract. 188, 135-140 (1992)

Fetal and Perinatal Infections A Consecutive Study H. Rudbeck R0ge and U. Henriques Aarhus University Hospital, Kommunehospitalet, Denmark

SUMMARY 118 cases of second trimester abortions, stillbirths, and perinatal deaths together with the placentas and case records were studied histologically and microbiologically to evaluate the incidence of infection and to assess possible correlations to certain clinical parameters. The infection rate was 39.4%, highest in the second trimester abortions (58.2%), where infection was also considered the most frequent cause of death (45.5%), though mostly without maternal signs of infection. 36.6% of cases with histological inflammation exhibited'positive culture results. The value of ordinary culture of the placenta and fetus is questionable, at least if not performed immediately after delivery. A significant correlation between infection and vaginal bleeding during pregnancy, ruptured membranes of more than 24 hours duration, and spontaneoHs labour was found. No protective effect of intact membranes was fOHnd.

Introduction

Prenatal and perinatal infections are still major problems often leading to spontaneous abortion or perinatal death. Only few studies have investigated both the fetus and the placenta as to chorioamnionitis and other signs of infection. The purpose of this study is to evaluate the incidence of infections in autopsies of second trimester abortions, stillbirths, and perinatal deaths. Ir attemps to extract possible correlations to certain clinical parameters not previously sufficiently investigated, such as amniocentesis, vaginal bleeding during pregnancy, placental hematoma shown by sonography, or intact membranes in relation to labour. The time of greatest risk of infection during pregnancy is estimated.

Material and Methods During the period January-lst 1987 to September 23rd 1988, 129 second trimester abortions, stillbirths, and perinatal deaths with placentas and case records were assessed by the University © 1992 by Gustav Fischer Verlag, Stuttgart

Institute of Pathology, Ärhus Kommunehospital. üf these, 11 cases were excluded because of severe maceration. The study material consists of 118 cases: 68 second trimester abortions (13-27 weeks of gestation), 31 stillbirths (28-42 weeks of gestation) and 19 perinatal deaths (26-41 weeks of gestation). 13 of the second trimester abortions and 1 of the perinatal deaths (34 weeks of gestation) were provoked abortionslinduced labour because of prenatally diagnosed severe malformations or on social indications. As there was no reason to suspect jnfection in these cases, they were treated as aseparate control group. In most cases, sterile culture from placenta, lungs, spinal fluid, liver, and intracardiac blood was undertaken to isolate aerobic and anaerobic bacteria and fungi. During the study period, unfortunately, we did not have the possibility to investigate for Mycoplasma hominis, Ureaplasma urealyticum or Clamydia trachomatis. To avoid contamination, iodine washing of the chorion was performed, followed by incision with a sterile knife and a swab placed in the placental tissue. The swab was placed in Stuarts medium and cultures were done in the University Department ofBacteriology, Ärhus Kommunehospital, according to routine methodology of that department. Po~itive culture from the fetus was defined as culture of bacteria from intracardiac blood, spinal fluid, or culture of the same species of bacteria from several sites. Immediately after delivery the fetus and placenta were stored in a refrigerator. The first morning after delivery a complete autopsy including examination of the placenta was performed. Histological ex am0344-0338/92/0188-0135$3.50/0

136 . H. Rudbeck IWge and U. Henriques ination of th e placenta, cord, and membranes as weil as of all fetal viscera including brain/meninges was undertaken. In a few ca ses only lung and brain/meninges were investigated. After fixation in formalin, the blocks were embedded in paraffin. The slides were stained with Hematoxylin-Giemsa 7 . All slides were evaluated by one of the authors (UH). As previous investigations had found that the overwhelming cause of inflammation is infectious 2, lj and as positive histology is better correlated to culture from amniotic fluid than to culture from the placenta itself4,26, infection is in this study defined as histological inflammation with or without positive culture. Chorioamnionitis is defined as a band of polymorphonuclear leucocytes in the chorion 4,5, 16,20,26 as shown in Fig, 1. Pneumonia is defined as polymorphonuclear leucocytes and fibrin in the alveolar spaces, interstitium or both 11, Sepsis is diagnosed by the presence of histological inflammation in association with positive culture from the same sites and from intracardiac blood, Information was obtained in th e case records about maternal age, number of pregnancies, maternal infectious disease in relation to abortion/delivery, vaginal bleeding during pregnancy, placental hematoma as shown by sonography, amniocentesis, duration of rupture of mem branes, and mode of delivery. For statistical evaluation the Mann Whitneys test was used,

Results The infection rates are shown in Table 1, More than half of the second trimester abortions exhibited infection. In nearly half of the cases it was considered the cause of death, as shown in Table 2. It was the most frequent cause of death in the second trimester abortions, In the stillbirths only about 10% had infection (as seen in Table 1), but in most cases, when present, it was considered the cause of death. The most frequent cause of death in this group was the mixed causes summarized in Table 2, In the perinatal deaths a little more than a quarter of the cases had infection and, when present, it was considered the cause of death. In the induced abortions/labours no signs of infection were found, and this group has been left out of the calculation of the results. The infectious diagnoses are shown in Table 3, Chorioamnionitis was the most important infection in both second trimester abortions (here often in combination with pneumonia), and in stillbirths, whereas sepsis was the dominating infection in the perinatal deaths. The number of ca ses in this group is, however, smalL T able 4 shows the relationship between histology and culture. 36.6% of cases with positive histology had positive culture, whereas only 9.1 % with negative histology had positive culture. Mainly enteric/vaginal bacteria were cultured as seen in Table 5. Table 1. Infection rates Infection rate

Infection as cause of death

39.4% (41/104)

31.7% (33/104)

Second trimester abortions

58.2% (32/55)

45.5 % (25/55)

Stillbirths

12.9% (4/31)

9.7% (3/31)

Perinatal deaths

27.8%

(5/18)

27.8% (5/18)

All ca ses

Induced abortions

0%

(0/14)

0%

(0/14)

Ta ble 2. Ca uses of death Infection Second trimester abortions

Others 1 Unknown

25

0

20

10

Stillbirths

3

3

15

10

Perina tal dea ths

5

5

8

0

Induced abortions

0

13

1

0

33

21

44

20

Total Fig, 1. A band of polymorphonuclear leucocytes in the chorion defining chorioamnionitis (x 25),

Severe malformations

1 Asphyxia, placental abruptio, placental infarct, intracranial bleeding, cervical insufficiency, induced abortion because of social indications,

Fetal and Perinatal Infections . 137

The me an maternal age was 30.2 years (range 17-41). 41 were primiparous, 77 were multiparous. There was no correlation benveen number of pregnancies and infection. 12 cases had infectious disease, e.g. pneumonia, none had signs of endometritis or amnionitis in relation to their abortion/delivery. In 11 of the cases infection of their placentalfetus was found. However, in 30 cases of infecti on no maternal infectious disease was found. The difference was not statistically significant. In 12 of the 104 cases an amniocentesis had been performed. In only 2 of these cases, infection was present and in neither case it was the cause of death. There was no positive statistical correlation benveen infection and amniocentesis. A significant correlation benveen infection and vaginal bleeding during pregnancy was found as shown in Table 6. A significant correlation benveen infection considered the cause of death and vaginal bleeding was also seen. Another investigated parameter was placental hematoma as shown by sonographic evaluation. Only 4 patients had a hematoma, 2 with infection and 2 without.

Table 5. Culture results Non-hemolytic streptococci Group B-streptococci E. coh Streptococcus faecalis Enterobacter cloacae Bacteroides group Pseudomonas group Klebsiella oxytoca Clostridium perfril1gel1s Acinetobacter Listeria monocytogenes Corynebacteria Staphylococcus albus Lactobacillus Staphylococcus aureus Citrobacter freundii Candida albical1s Yeasts

4 2 3 3 3 3 1 2 1 2 1 1 3 2 1 1 2 1

Total

36

Table 6 Table 3. Infectious diagnoses (number of ca ses with infection as cause of death in brackets) Second tri: Stillbirths Perinatal mester deaths

14

36

- Vaginal bleeding

19

49

68

Total

41

63

104

21 (18)

Chorioamnionitis 12 (8) and pneumonia

0(0)

0(0)

12 (8)

1 (1)

0(0)

0(0)

1 (1)

Sepsis

1 (1)

0(0)

4 (4)

5 (5)

Others (1 CMV, 1 bronchitis)

2 (1)

0(0)

0(0)

2 (1)

32 (25)

4 (3)

5 (5)

41 (33)

Table 4. Histology versus culture Total Positive histology Positive culture

15

Positive histology Negative culture

19

Infection as cause of death versus vaginal bleeding during pregnancy Infection as Other causes cause of death of deaths

7 41

Negative histology Positive culture

7

29

Total p = 0.05 1

+ Vaginal bleeding

16

20

36

- Vaginal bleeding

17

51

68

Total

33

71

104

1

Negative histology Negative culture

Negative histology Culture not undertaken

Infection versus vaginal bleeding during pregnancy 22

1 (1)

Positive histology Culture not undertaken

Total p = 0.003 1

+ Vaginal bleeding

4 (3)

Total ca ses

- Infection

Total ca ses

Chorioamnionitis 16 (14)

Pneumonia

+ Infection

Mann-\1Vhimeys test.

Table 7 shows the relationship benveen infection and intact membranes, defined as delivery in membranes, or delivery within 10 minutes after rupture of membranes. There are as many cases in which infection was associated with ihtact membranes as there are cases without such an association. The same pattern is seen where infection is considered the cause of death. Conversely, as shown in Table 8, there is a significant correlation benveen rupture of membranes of more than

138 . H. Rudbeck fuJge and U. Henriques

24 ho urs duration and irifection, - but not when infection is considered the cause of death. Table 9 shows infection versus spontaneous/provoked labour. There is a statistical correlation between spontaneous delivery and infection, both in general and where infection is considered the cause of death.

Infection as cause of death versus rupture of membranes of more than 24 hours duration

7

8

13

Tablc 7

Rupture of 26 membranes < 24 hours duration

65

91

Total

71

104

- Infection

Total p = 0.00]2

+ Infection

- Infection

Total NS2 (p = 0.17)

Infection versus intact membranes 1 Intact mem- 11 branes

11

22

Rupture of membranes > 10 minutes

30

52

82

Total

41

Other causes Infection as cause of death of death Rupture of membran es > 24 hours duration

1

33

Total NSI

(p = 0.07)

Mann-Whitneys test.

Table 9

+ Infection

104

63

Infection versus spontaneous/provoked labour Infection as cause of death versus intact

membranes l

Infection as Other causes cause of death of death

Total NS2 (p = 0.11)

Intact mem- 10 branes

12

22

Rupture of membranes > 10 minutes

23

59

82

Total

33

Spontaneous 31 labour

26

57

Provoked la- 9 bour

37

46

40

63

1031

Total

Infection as cause of death versus spontaneous/provoked labour

71

Infection as Other causes cause of death of death

104

1 Infant delivered in membranes or delivery within 10 minutes after rupture of membranes. - 2 Mann-Whitneys test.

Table 8

Spontaneous 27 labour

30

57

Provoked labour

6

40

46

33

70

Total

+ Infection

- Infection

Total p = 0.0013 1

Infection versus rupture of membranes of more than 24 ho urs duration Rupture of membranes >24hours duration

11

2

Rupture of membranes < 24hours duration

30

61

91

Total

41

63

104

13

Total p = 0.00]2

1

1 case not stated. - 2 Mann-Whitneys test.

Discussion This study confirms that chorioamnionitis, often in combination with pneumonia, is a frequent cause of fetal death, especially so in the se co nd trimester, and mostly as a silent chorioamnionitis without maternal signs of infection. This is in agreement with earlier studies, although these did not investigate risk time during pregnancy. In Naeye's series J J, 12 infection was among the most frequent causes of neonatal deaths, and other studies have also revealed the significant role played by infection J7- J9 , although not as important in late intrauterine death 21 .

Fetal and Perinatal Infections . 139

Group B streptococci have been singled out as responsible for reproductive lossi, 2, 9 especially in midgestation 24 . In our study we had 2 cases of infection with group B streptococci. Anothe,r important causative agent is Listeria monocytogenes I ,2,17, which in our study was responsible for 1 case. However, we identified the microbial agent only in 36.6% of cases. Other investigators have suffered from the same difficulties in identifying the microbial agents and have found the value of culturing doubtfu14, 25. Many more re cent studies have demonstrated a high incidence of Mycoplasma hominis I ,2,4,1O, Ureaplasma urealyticum I , 8, 18, 19 and maybe Clamydia trachomatis l . It seems possible that many of our negative cultures might have had one of these microbials as the agent. However, of the cultured bacteria all but one are entericlvaginal, indicating an ascending infection. Other causes of inflammation in the placenta and membranes have been postulated, such as changes in pH, anoxia or meconium. The conclusion has been reached that the overwhelming cause of inflammation is infection, most frequently ascending2, 15. In our study the "control group" consisting of induced abortions/labours in no instance had signs of inflammation, supporting this assertion. Contamination of the placenta is unavoidable as it passes the birth canal. However, the growth of these bacteria was minimized as the placentas were stored in a refrigerator immediately after delivery. Still, contamination must be responsible for the positive cultures in absence of inflammation. In our study we found a significant correlation between vaginal bleeding and infection. Of course, we could not determine whether the bleeding was a result of infection or vice versa. An earlier study claimed a correlation between infection with Mycoplasma hominis and threatening abortion lO and others have also found abnormal bleeding in mothers with intrauterine infection3,Il. We did not find any correlation to placental hematoma as visualised by sonographic evaluation. We have not succeeded in finding other references evaluating this parameter. Amniocentesis is a possible source of infection. This is, however, a minor problem. In our study we had 2 women with placental infection where amniocentesis was undertaken. In neither ca se was it the cause of death. Another study states the incidence of chorioamnionitis after amniocentesis to be 0.19-0.71 %23. From our da ta it seems that intact membranes are no protection against infection. This fact, defined as the amnion infection syndrome I2 , is confirmed by others 2,3,9,13. It seems that under unfavorable conditions, e.g. cervical insufficiency (but also with a closed cervical os), bacteria can force through the barrier of the membranes and cause a local chorioamnionitis which may spread and become universall, 13. Chorioamnionitis is a well-established factor in premature labour2 ,6, 14, 16, 19, 24, 2S., most likely acting, by activating prostagiandin production 1. There is also a correlation between presence of acute inflammation and occurrence of

labour at term 22 . Concurrent to this we found a highly significant correlation between infection and spontaneous labour. Silent chorioamnionitis is rare in normal pregnancy22. This is in agreement with our study, in which our "control group" revealed no instance of inflammation. The well-established fact of increased incidence of infection with prolonged rupture of membranes 2 , 16, 19 was also no ted in our study. However, infection may be the etiology of rupture of membranes thereby spreading to the rest of the amnionic cavity and the fetus. In conclusion, infection is frequent and is a major cause of death, especially so in the second trimester and mostly without maternal signs of infection. We found a significant correlation between infection and vaginal bleeding during pregnancy; ruptured membranes of more than 24 hours duration; and spontaneous labour. We found no protection by intact membranes. The value of ordinary culture of placenta and fetus is questionable, at least if not performed immediately after delivery .... References 1 Benirsehke K, Robb JA (1987) Infeetious eauses of fetal death. Clin Obstet Gyneeol 30: 284-294 2 Blanc WA (1981) Pathology of the placenta, membranes and umbilieal cord in baeterial, fungal and viral infeetions in man. In: Perinatal Oiseases. Monographs in Pathology 22: 67-132. Baltimore 3 Oesa OJ, Trevenen Cl (1984) Intrauterine infections with group B beta-hemolytie streptocci. Br J Obstet Gynecol 91: 237-239 4 Oong Y, St. Clair PJ, Ramzy I, Kagan-Hallet KS, Gibbs RS (1987) A microbiologic and clinieal study of placental inflammation at term. Obstct Gynecol 70: 175-182 5 Fox H, Langley FA (1971) Leucoeytic infiltration of the placenta and umbilical cord. A clinico-pathological study. Obstet Gynecol 37: 451-458 '6 Guzick OS, Winn K (1985) The association of chorioamnionitis with preterm delivery. Obstet Gynecol 65: 11-16 7 Henriques U (1970) Hematoxylin-Giemsa. An improved stain for routine diagnostic histology and cytology. Acta Path Micr Scand A, 78: 236-237 8 Hillier SL, Martius J, Krohn M, Kiviat N, Holmes KK, Esehenbach DA (1988) A case-control study of chorioamniotic infection and histologie chorioamnionitis in prematurity. N EnglJ Med 319: 972-978 9 Katz V, Bowes WA (1988) Perinatal group B streptoccal infections across intact amniotic membranes. J Reprod Med 33: 445--449 10 Munday PE, Porter R, Falder PF et al. (1984) Spontaneous abortion an infectious aetiology? Br J Obstet Gynecol 91: 1177-1180 11 Naeye RL, Oellinger WS, Blane WA (1971) Fetal and maternal features of antenatal bacterial infections. J Pediatr 79: 733-739 12 Naeye RL (1977) Causes of perinatal mortality in the US collaborative perinatal project. JAMA 238: 228-229 13 Naeye RL, Peters EC (1978) Amniotic fluid infections with intaet membranes leading to perinatal death: A prospective study. Pediatrics 61: 171-177 14 Naeye RL (1982) Factors that predispose to premature rupture of the fetal membranes. Obstet Gynecol 60: 93-98

140 . H. Rudbeck Iillge and U. Henriques 15 Pankuch GA, Appelbaum PC, Lorenz RP, BottiJJ, Schachter ], Naeye RL (1984) Placental microbiology and histology and the pathogenesis of chorioamnionitis. Obstet Gynecol 64: 802-806 16 Perkins RP, Zhou S-M, Butler C, Skipper B] (1987) Histologie chorioamnionitis in pregnancies of various gestational age: Irriplications in preterm rupture of membranes. Obstet Gynecol 70: 856-860 17 Pryse-Davies ], Hurley R (1979) Infections and perinatal mortality. ] Antimicrob Chemother 5A: 59-70 18 Quinn PA, Butany ], Chipman M, Taylor ], Hannah W (1985) A prospective study of microbial infection in stillbirths and early neonatal death. Am] Obstet Gynecol151: 238-249 19 Quinn PA, Butany], Taylor], Hannah W (1987) Chorioamnionitis: Its association with pregnancy outcome and microbial infection. Am J Obstet Gynecol156: 379-387 20 Rayburn W, Sander C, Barr M, Rygiel R (1985) The stillborn fetus: Placental histologie examination in determining a cause. Obstet Gynecol 65: 637-641

21 Royston D, Geoghegan F (1985) Amniotic fluid infection with intact membranes in relation to stillborns. Obstet Gynecol 65:745-746 22 Salafia CM, Weigl C, Silberman L (1989) The prevalence and distribution of acute placental inflammation in uncomplicated term pregnancies. Obstet Gynecol 73: 383-389 23 Siekmann U, Daschner F, Heilmann L (1985) Chorioamniale Infektionen nach diagnostischer Amniocentese im Ir. Trimenon. Z Geburtsch u. Perinat 189: 119-124 24 Singer DB, Campognone P (1986) Perinatal group B streptococcal infection in midgestation. Pediatr Pathol 5: 271-276 25 Svensson L, Ingemarsson I, Mardh P-A (1986) Chorioamnionitis and the isolation of microorganisms from the placenta. Obstet Gynecol 67: 403-409 26 Zhang], Kraus FT, Aquino TI (1985) Chorioamnionitis: a comparative histologie, bacteriologic and clinical study. Int ] Gynecol Pathol 4: 1-10

Received November 12, 1990 . Accepted in revised form April 8, 1991

Key words: Chorioamnionitis - Fetal death - Fetal membranes - Pregnancy complications - Infections Hanne Rudbeck Rage, Institute of Pathology, Aarhus University Hospital, Kommunehospitalet, DK-8000 Aarhus, Denmark