Abstract
Objective
The study was designed to examine the hypothesis whether the course and severity of hepatitis E virus (HEV)–related liver disease is worse during pregnancy.
Method
The prospective study included 1088 patients (550 pregnant; 538 nonpregnant) with clinically and biochemically confirmed acute viral hepatitis (AVH) or acute liver failure (ALF) and were subjected to a complete panel of hepatitis serology.
Results
In the pregnant cohort, HEV was the cause of infection in 80.36% (442/550) of cases, whereas non-HEV accounted for 19.63 (108/550) of cases. In the ALF pregnant group, the prevalence of HEV was observed in 73.38% (102/139) of cases, whereas other viruses accounted for 26.61% (37/139) of illness. Ninety-eight of 129 (75.96%) cases of HEV-infected pregnant women died, whereas non-HEV infection was responsible for only 31 of 129 (24.04%) cases' death in comparison. Serum viral load in the ALF group was also significantly higher than that in the AVH group in the pregnant (24578.6 ± 12410.3 vs. 6821.9 ± 1832.7, respectively) cohort and nonpregnant cohort (583.6 ± 187.34 vs. 298.68 ± 65.77, respectively).
Conclusion
HEV infection has a higher incidence, more severe course, and greater mortality in the pregnant cohort than in the nonpregnant cohort.
Keywords: acute liver failure, acute viral hepatitis, hepatitis E virus, pregnancy
Abbreviations: ALF, Acute Liver Failure; AVH, Acute Viral Hepatitis; FHF, Fulminant Hepatic Failure; HEV, Hepatitis E Virus; PCR, Polymerase Chain Reaction; RNA, Ribonucleic Acid
Hepatitis E virus (HEV) is a major cause of acute viral hepatitis (AVH) in developing countries including India. Predicting definitive outcomes related to HEV infection during pregnancy has been marred with contradictory results from different studies. There have been numerous studies associating pregnancy with poor maternal and fetal outcomes from HEV-related infections.1, 2, 3, 4 Pregnancy by itself does not seem to predispose individuals to HEV infection, but it is a risk factor for viral replication and predispose individuals to developing fulminant hepatitis.2,5,6 However, recently, a study from a large tertiary care hospital in North India failed to appreciate any difference in mortality related to either pregnancy status or HEV as etiological agents.7 Malnutrition, high prevalence of folate deficiency, steroid hormones, and inversion of T and B lymphocytes in early pregnancy have been postulated to be the contributing factors.6 Mortality rates among pregnant women infected with HEV have been reported to be nearly 25% with higher rates between 40 and 60% resulting from acute liver failure (ALF) caused by HEV infection.4,7,8 A significant proportion of pregnant women with acute hepatitis E (up to 70%) progress to ALF with a short pre-encephalopathy period, the rapid development of cerebral edema, and frequent occurrence of disseminated intravascular coagulation.9 Mother to child transmission of HEV has been reported to be 33%, 50%, and even 100% in different studies.8,10,11 The fatality rate among pregnant women with ALF is reported to be high at 22.2%, with the maximum severity occurring during the third trimester (44.4%).11,12 HEV infection during pregnancy is also associated with high rates of spontaneous abortion, intrauterine death, and preterm labor.8,11 The primary objective of this study was to compare maternal outcomes of HEV-related liver disease with those of non–HEV-related liver disease. The secondary objective was to investigate and compare clinical and biochemical parameters in AVH and ALF in the pregnant and nonpregnant population.
Materials and methods
The prospective clinical study was conducted in 1088 female patients of reproductive age group between 15 and 45 years of age with confirmed hepatitis viral illness recruited from the Department of Obstetrics and Gynecology, Lok Nayak Hospital, New Delhi, and those presenting in the Outpatient and Emergency Department, Department of Internal Medicine, Lok Nayak Hospital, New Delhi, over a period of 5 years between 2005 and 2010. Of 1088 patients, 768 patients had AVH and 320 patients presented with ALF. AVH was defined as an acute illness with the discrete onset of symptoms and jaundice or elevated serum aminotransferase levels.13 Diagnosis of ALF was made when the patient, with no known pre-existing liver disease, had hepatic encephalopathy within 4 weeks of the onset of disease.14 A total of 550 of 1088 patients represented the pregnant cohort in all three trimesters of pregnancy, whereas 538 subjects in the study were not pregnant. Pregnancy status of patients was confirmed before recruiting them into either of the cohorts. The pregnant cohort (550) included 411 patients with AVH and 139 patients with ALF, whereas the nonpregnant cohort (538) included 357 patients with AVH and 181 patients with ALF. A written informed consent was obtained from all the patients.
The study was approved by the Institutional Ethics Committee of the Maulana Azad Medical College, New Delhi. Serum samples were collected and stored at −70 °C, and serological tests were performed for viral markers. Viral RNA extraction was performed for those patients who were found to be positive for anti–HEV IgM from 140 μl of serum using a viral RNA extraction kit (QIAamp Viral RNA Mini Kit 52904). HEV RNA load (copies/ml) was determined for those AVH and ALF samples which were positive for HEV RNA using a real-time PCR machine (Rotor–Gene bRG-2000; Corbett Research) using Geno Sen's HEV Real Time PCR Reagents kit, India. The representative amplified products were purified, sequenced commercially, and compared with the HEV sequences in the Gen Bank using the CLUSTAL W analysis software, and the phylogenetic tree was constructed using the PHYLIP package software using the neighbor-joining method based on the partial nucleotide sequence of the Open reading frame (ORF1) region. The student t-test was used to analyze mean ± standard deviation in both independent and dependent samples. The Fisher exact test was used for analyzing the frequency data. The Mann-Whitney test was used to analyze the viral load. The calculations were made using the Epi Info, version 6, software of the Atlanta Center for Disease Control. The differences were considered significant for P values less than 0.05.
Results
HEV was the most prevalent hepatotropic virus in the study groups and was responsible for 80.36% (442/550) of cases in the pregnant cohort. Clinical and biochemical parameters of AVH and ALF in the pregnant and nonpregnant states are compared in Table 1. HEV infection was observed in 82.72% (340/411) of cases of AVH pregnant group but in only 43.41% (155/357) of cases in the AVH nonpregnant group. In the ALF cases, the prevalence of HEV was observed in 73.38% (102/139) of cases in the pregnant group and in 61.32% (111/181) of cases in the nonpregnant group. Prevalence data of hepatotropic viruses in AVH and ALF in pregnant and nonpregnant states are compared in Table 2, Table 3, respectively.
Table 1.
The Comparative Analysis of Biochemical and Clinical Parameters in Patients With AVH and ALF.
| Parameters | AVH pregnant (411) | AVH nonpregnant (357) | P value | ALF pregnant (139) | ALF nonpregnant (181) | P value |
|---|---|---|---|---|---|---|
| Hb | 9.30 ± 3.11 | 11.21 ± 1.91 | <0.001 | 10.42 ± 2.02 | 8.74 ± 2.21 | <0.001 |
| ALT IU/dl | 471.18 ± 369.85 | 486.62 ± 502.81 | 0.62 | 806.31 ± 574.91 | 1124.00 ± 458.00 | <0.001 |
| AST IU/dl | 557.82 ± 421.63 | 545.43 ± 615.64 | 0.74 | 663.15 ± 351.77 | 803.00 ± 375.00 | <0.001 |
| TBil | 6.90 ± 2.32 | 5.70 ± 2.41 | <0.001 | 11.75 ± 3.82 | 13.62 ± 7.34 | <0.001 |
| ALP | 156.81 ± 142.82 | 187.23 ± 122.16 | <0.001 | 247.5 ± 262.3 | 296.3 ± 322.3 | <0.14 |
| Fever | 319 (77.6%) | 197 (55.2%) | <0.001 | 124 (89.2%) | 64 (35.3%) | <0.001 |
| Pruritus | 140 (34.1%) | 153 (42.9%) | 0.15 | 82 (59%) | 95 (52.5%) | 0.29 |
| Anorexia | 389 (94.6%) | 313 (87.7%) | <0.001 | 139 (100%) | 128 (70.7%) | <0.001 |
| Pedal edema | 335 (81.5%) | 110 (30.8%) | <0.001 | 117 (84.2%) | 85 (46.9%) | <0.001 |
| IUD | 103 (25.1%) | NA | NA | 120 (86.33%) | NA | NA |
| Maternal mortality | 13 (3.16%) | 0 | <0.001 | 116 (83.45%) | 30 (16.6%) | <0.001 |
| Preterm delivery | 173 (42.1%) | NA | NA | 102 (73.4%) | NA | NA |
AVH, acute viral hepatitis; ALF, acute liver failure; ALT, alanine aminotransferase; AST, aspartate aminotransferase; TBil, total bilirubin; ALP, alkaline phosphatase; IUD, intrauterine death; NA, non applicable; hb, hepatitis B.
Table 2.
Prevalence of Hepatotropic Viruses in AVH.
| Viral markers | AVH pregnant (411) | AVH nonpregnant (357) | P value |
|---|---|---|---|
| HAV IgM | 13 (3.16%) | 44 (12.32%) | <0.001 |
| HBsAg | 39 (9.48%) | 77 (21.56%) | <0.001 |
| Anti-HCV | 11 (2.67%) | 7 (1.96%) | 0.67 |
| HEV IgM | 340 (82.72%) | 155 (43.41%) | <0.001 |
| Non–A-E | 8 (1.94%) | 74 (20.72%) | <0.001 |
AVH, acute viral hepatitis; HAV IgM, hepatitis A virus immunoglobulin M; HBsAg, hepatitis B′s antigen; HCV, hepatitis C virus; HEV IgM, hepatitis E virus immunoglobulin M; non–A-E, non–adverse events.
Table 3.
Prevalence of Hepatotropic Viruses in ALF.
| Viral markers | ALF pregnant (139) | ALF nonpregnant (181) | P value |
|---|---|---|---|
| HAV IgM | 5 (3.6%) | 5 (2.76%) | 0.91 |
| HBsAg | 13 (9.35%) | 28 (15.47%) | 0.14 |
| Anti-HCV | 1 (0.72%) | 2 (1.10%) | 0.81 |
| HEV IgM | 102 (73.38%) | 111 (61.32%) | 0.03 |
| Non–A-E | 18 (12.94%) | 35 (19.34%) | 0.17 |
ALF, acute liver failure; HAV IgM, hepatitis A virus immunoglobulin M; HBsAg, hepatitis B′s antigen; HCV, hepatitis C virus; HEV IgM, hepatitis E virus immunoglobulin M; non–A-E, non–adverse events.
HEV RNA was present in 39.12% (277/708) of the HEV IgM–positive cases including both pregnant and nonpregnant groups. HEV RNA was present in 42.30% (187/442) of HEV IgM–positive cases in the pregnant group, whereas in the nonpregnant group, it was present in only 33.8% (90/266) cases. HEV RNA was detected in the serum in 79.41% of cases (81/102) of ALF, whereas in the AVH group, it was detected in only 31.17% (106/340) of cases.
Serum viral load in the ALF group was also significantly higher than that in the AVH group in the pregnant (24578.6 ± 12410.3 copies/ml vs. 6821.9 ± 1832.7 copies/ml, P < 0.05) cohort and nonpregnant cohort (583.6 ± 187.34 copies/ml vs. 298.68 ± 65.77 copies/ml, P < 0.05). Genotyping of representative HEV samples showed that all the cases belonged to genotype 1.
Of the 139 pregnant patients with ALF, the maternal mortality was recorded in 83.45% (116/139) of cases, whereas only 3.16% (13/411) of cases in the pregnant AVH cohort died. The most common etiological agent responsible for mortality in the ALF pregnant cohort and AVH pregnant cohort was HEV, responsible for 76.72% (89/116) and 69.23% (9/13) of cases, respectively. Twenty-two of 89 patients (24.7%) were in the second trimester while the remaining 67 patients (75.2%) were in the third trimester. Of the 9 patients who died due to HEV-related AVH, 2 (22.22%) patients were in the second trimester while the remaining 7 cases (77.77%) were in the third trimester. Mortality data with respect to different viruses are presented in Table 4.
Table 4.
Etiology of Maternal Mortality in AVH and ALF Pregnant Cases.
| Viral markers | AVH13 | ALF (116) | P value |
|---|---|---|---|
| HAV | 1 (7.69%) | 5 (4.31%) | 0.88 |
| HBV | 2 (15.38%) | 7 (6.03%) | 0.49 |
| HCV | 0 (0%) | 1 (0.86%) | 0.18 |
| HEV | 9 (69.23%) | 89 (76.72%) | 0.79 |
| Non–A-E | 1 (7.69%) | 14 (12.06%) | 0.99 |
AVH, acute viral hepatitis; ALF, acute liver failure; HAV, hepatitis A virus; HBV, hepatitis B virus; HCV, hepatitis C virus; HEV, hepatitis E virus.
Fetal mortality was observed in 86.33% (120/139) of cases of pregnancy complicated by ALF and in 25.06% (103/411) of cases of pregnancy complicated by AVH. Patients with ALF mostly had preterm delivery which was observed in 73.33% (102/139) of cases, and only 42% (173/411) of pregnant women with AVH delivered prematurely.
While comparing the outcomes of the HEV-related and non–HEV-related cases, it was observed that the mortality rate was much higher in HEV-related cases than in the non-HEV group. Ninety-eight of 129 cases (75.96%) of HEV-infected pregnant women died, whereas non-HEV was responsible for only 24.04% (31/129) cases’ death in comparison (P < 0.001). Comparison between outcomes of HEV and non–HEV-related cases is presented in Table 5.
Table 5.
Comparison Between HEV and Non–HEV-Associated MMortality.
| Pregnancy mortality (129) | ||||
|---|---|---|---|---|
| AVH13 | ALF (116) | Total | P value | |
| HEV | 9 | 89 | 98 | <0.001 |
| Non-HEV | 4 | 27 | 31 | |
HEV, hepatitis E virus; AVH, acute viral hepatitis; ALF, acute liver failure.
Discussion
HEV infection is one of the predominant causes of pregnancy-related complications in developing countries including India.2 HEV alone was responsible for 80.36% of cases of hepatitis virus infection among pregnant women in our study. The mortality among ALF cases in our study was more confined to the HEV-infected cohort than to the other hepatotropic viruses–infected cohort. In the present study, HEV infection during pregnancy resulted in fetal and/or maternal mortality, abortion, premature delivery, or death of a live-born baby soon after birth which validates the notion that pregnancy is a risk factor for viral replication and, subsequently, a more severe form of the disease. The mortality rate was also found to be much higher in the pregnant group than in the nonpregnant group in our study. Hence, pregnancy is a risk factor in women with HEV-related ALF for more severe disease and subsequently poorer outcomes in contrast to a previous study in the same region.7 Various studies from different regions of the Indian subcontinent report prevalence of HEV in AVH during pregnancy between 58 and 86%.4,12,15 In another study, pregnant women infected with HEV had a worse outcome than women infected with other viruses.3 High levels of steroid hormones during pregnancy are immunosuppressive, promote viral replication, and also have a direct inhibition on hepatic cells, which may predispose individuals to hepatic dysfunction/failure, higher viral loads, and subsequently poorer outcomes when exposed to infectious pathogens.16, 17, 18 We tried to compare viral load between the pregnant and nonpregnant group in this study, and it was found to be significantly higher in the pregnant group than in the nonpregnant group. In a previous study by Kar et al.,19 a comparatively higher HEV viral load was observed in patients with fulminant hepatic failure (FHF) than in patients with AVH in both the pregnant and nonpregnant groups. There are differences in the maternal and fetal outcome of pregnancy associated with viral hepatitis in various studies in different geographical regions.20 Increased maternal and fetal mortality has been reported in many studies, mainly from developing countries. The liver injury due to HEV infection often takes a severe course during pregnancy, and it needs to be studied in more detail whether the severity of the infection is regulated by immune system changes, hormonal factors, and genetic and host factors. Despite limitations in our study, our results substantiate that when HEV infection occurs during pregnancy, the course of ALF is stormy and carries a high mortality.
Authors' contribution
R.K. contributed to data collection, article writing, and proofreading of article. R.K.H. contributed to statistical analysis and article writing. J.B. contributed to statistical analysis and article writing. A.K. contributed to conceptualization, guidance, supervision, and protocol development. P.K. contributed to conceptualization, guidance, supervision, and protocol development.
Conflicts of interest
The authors have none to declare.
Acknowledgement
The authors acknowledge the efforts of all participants who gave consent to be a part of the study.
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