Clinical Outcomes and Prevalence of Hepatitis E Virus (HEV) Among Non-A-C Hepatitis Patients in Egypt

Figures & data

Figure 1 Schematic flow of the study design. Acute hepatitis symptomatic patients admitted to Assiut University hospitals were assessed for LFTs including liver transaminases and bilirubin. Patients with abnormal LFTs were screened for routine viral hepatitis markers (HAV, HBV, HCV, CMV, and EBV), autoimmune hepatitis markers. Medical history for the patients excluded the possibility of DILI or alcohol-induced liver injury. Samples tested negative to the previously mentioned markers were screened for HEV markers. Patients with liver disease of known etiology (for example HBV, HCV, autoimmune hepatitis, and DILI were not included in this study.

Figure 2 Assessment of HEV markers in AHUE and the outcomes of infection. AHUE (n=300) were screened for HEV markers (anti-HEV IgM, anti-HEV IgG, and HEV RNA). 30 samples were reactive to HEV markers. Four samples were positive to the tested HEV markers (Group 1), 3 samples were positive to anti-HEV IgG and HEV RNA (Group 5) and 2 samples tested positive for anti-HEV IgG (group 6). All the previous patients (Group 1, 5 and 6) recovered without complications. 8 samples were positive to anti-HEV IgM and HEV RNA, and negative for anti-HEV IgG (Group 2), 4 samples tested positive to anti-HEV IgM and anti-HEV IgG, and negative for HEV RNA (Group 3), and 9 samples were tested positive for anti-HEV IgM (Group 4). The previous patients (Group 2, 3, 4, n= 21), 17 patients recovered without complications and 4 patients progressed to FHF. Groups 4 and 6 were also positive for HEV Ag.

Table 1 Demographic and Laboratory of AHE and NHE Infected Patients

Variables	AHE Patients (n=30)	NHE Patients (n=270)	Statistic S/Ns^a
Age (year)	51 (44–60)	49 (40–56)	Ns
Sex M/F	16/14 (53%/ 47%)	130/140 (48%/ 52%)	Ns
ALT U/l	454 (236–946)	183 (99–412)	S, P < 0.0001
AST U/l	357 (221–630)	134 (89–224)	S, P < 0.0001
ALP U/l	300 (173–412)	321 (217–495)	Ns
ALT/ALP (R-value)^b	5.14 (3.2–5.8)	1.89 (1.386–2.76)	S, P < 0.0001
Bilirubin (µmol/L)	240 (140–260)	243 (156–320)	Ns

Notes: All values are represented as medians and interquartile ranges (IQR). ALT (normal range < 40 U/l); AST (normal range < 35 U/l) and ALP (normal range, 35–105 U/l); bilirubin (normal range, 1.71 to 20.5 μmol/l). ^aS means significant (p<0.05) and Ns means non-significant (p>0.05) as calculated by student’s unpaired t test. ^bR-value= (ALT/ULN)/ (ALP/ULN). ULN is the upper limit normal.

Table 2 Baseline Characteristics of Patients Developed Fulminant Hepatic Failure (FHF)

Parameters	Patient 1	Patient 2	Patient 3	Patient 4
Age (years)	60	72	66	76
Sex (M/F)	M	M	M	F
Liver Function tests	ALT 1025 U/l	ALT 845 U/l	ALT 1220 U/l	ALT 459 U/l
	AST 683 U/l	AST 563 U/l	AST 813 U/l	AST 306 U/l
	ALP 520 U/l	ALP 340 U/l	ALP 410 U/l	ALP 178 U/l
	Bilirubin 480 µmol/L	Bilirubin 520 µmol/L	Bilirubin 640 µmol/L	Bilirubin 450 µmol/L
International normalized ratio (INR)	1.55	1.7	1.8	1.67
HEV markers	Anti-HEV IgM (+), HEV RNA (+), Anti-HEV IgG (-).	Anti-HEV IgM (+), HEV RNA (+), Anti-HEV IgG (-).	Anti-HEV IgM (+), Anti-HEV IgG (+), HEV RNA (-).	Anti-HEV IgM (+), Anti-HEV IgG (-), HEV RNA (-).
HEV genotype/subtype	HEV-1 1e	HEV-1 1b	ND	ND
History of liver disease	Yes	Yes	No	No
Malignancy	No	No	Yes Leukemia	No
Hospitalization	22 days	33 days	37 days	26 days
Rural/Urban communities	Rural	Rural	Rural	Rural

Table 3 Comparison of Characteristics of FHF Patients and Acute Self-Limited HEV Patients

Parameters	FHF Patients (n=4)	Self-Limited AHE Patient (n=26)	Statistic S/Ns^a
Age (years)	69 (61–75)	51 (45–61)	S, p= 0.0472
Sex (M/F)	3/1	13/13	Ns
Liver function tests (LFTs)	ALT 935 (555–1171)	ALT 430 (223–873)	S, p=0.0467
	AST 623 (370–780)	AST 315 (189–539)	S, p=0.0486
	ALP 375 (218–492)	ALP 285 (157–379)	Ns
	Bilirubin 500 (457–610)	Bilirubin 230 (109–250)	S, p< 0.0001
INR	1.685 (1.58–1.775)	1.24(1.19–1.315)	S, p< 0.0001
Complications coagulopathy, encephalopathy, and death	Yes (4/4)	No (0/26)	S, p< 0.0001
HEV markers	Anti-HEV IgM 4/4 (100%) Anti-HEV IgG 1/4 (25%) HEV RNA 2/4 (50%)	Anti-HEV IgM 21/26 (80.7%) Anti-HEV IgG 12/26 (46%) HEV RNA 13/26 (50%)	NA
HEV genotype/subtype and mean viral load	HEV-1 (subtype 1b, 1e) 6.8x 10^3 IU/mL	HEV-1 (subtype 1b,1e), and HEV-3 (subtype 3a) 2.76x 10^3 IU/mL	Ns
History of liver disease	2/4 (50%)	5/26 (19.2%)	S, p=0.0356
Hospitalization (days)	29 (23–36)	7 (5–10)	S, p< 0.0001
Rural/Urban communities	Rural (4/4) Urban (0/4)	Rural (18/26) Urban (8/26)	Ns

Notes: The values are represented as medians and interquartile ranges. ^aS means significant (p<0.05) and Ns means non-significant (p>0.05) as calculated by student’s unpaired t test.

Figure 3 Comparison of LFTs and HEV markers in FHF patients and self-limited AHE patients. FHF patients (n=4) and self-limited AHE patients (n=26) were assessed for LFTs and HEV markers. (A) The LFTs (ALT, AST, and ALP) were measured in FHF patients and self-limited AHE patients. (B) The absorbance of anti-HEV IgM (A₄₅₀/C.O.) was compared in FHF patients and self-limited AHE patients. *, ** means p<0.05 and p≤ 0.01, respectively as determined by two-tailed student’s t test.

Figure 4 Phylogenetic analysis of the isolated viruses. A maximum-likelihood phylogenetic tree was constructed using various HEV reference sequences with 1000 bootstrap replicas. GenBank accession numbers are shown for each HEV reference strain used in the phylogenetic analysis. The isolated viruses from the patients were remarked it by putting a circle beside them.