journalof Viral Hepatitis, 1996. 3 ,20 3-205

Hepatitis E virus infection in patients from Saudi Arabia with sickle cell anaemia and P-thalassemia major: possible transmission by blood transfusion I, Al-Fawaz,' S. Al-Rasheed, M. Al-Mugeiren,' A. Al-Salloum, M. Al-Sohaibani' and S. Ramia' Departments oJ'Pediatrics and 2Pathology. College oJMedicine and King Khalid University Hospital, King Saud University, Riyadh, Saudi Arabia

Received 9 February 9Y6; accepted for publication 25 March 19 96

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SUMMARY. The seroprevalence of antibodies against hepatitis E virus (HEV) and hepatitis C virus (HCV) was investigated in Saudi children with sickle cell anaemia (SCA) (50 patients: 28 boys, 22 girls: age range 2-14 years) and P-thalassemia major (28 patients: 12 boys, 16 girls: age range 2-12 years). The SCA patients were from the Gizan area (South) while the thalassemics were from the Riyadh area (Central province). The prevalence of hepatitis E virus antibody (HEVAb) in patients with SCA (18.0%) and in those with p-tha- lassemia major (10.7%) was higher than in the control groups (5.5% and 2.8%) but this did not reach the level of statistical significance. In contrast to the situation with HEV, hepatitis C virus antibody (HCVAb) positiv-

ity was significantly higher in patients with SCA (16.0%) and in thalassemics (57.1%) than in the respective control groups. Although the difference in HEV seropositivity between P-thalassemia major, SCA patients and their respective controls is not statistically significant, the possibility of blood-borne HEV in the Saudi population cannot be excluded. Further investi- gations using HEV-specific polymerase chain reaction techniques are required to confirm whether transmis- sion of HEV through blood preparations or transfusion is possible.

Keywords: HEV, sickle cell anaemia, P-thalassemia major, blood transfusion.

INTRODUCTION

There are at least two types of non-A, non-B hepatitis. One is transmitted parenterally and is caused by the hepatitis C virus (HCV) [l], and the other is transmitted enterally and is caused by the hepatitis E virus (HEV) [2,3]. Hepatitis caused by HEV occurs in both epidemic and sporadic forms-epidemics generally occur in developing countries and are mainly the result of con- taminated water supplies [4-61. Sporadic cases, how- ever, have been reported in developed countries among travellers returning from endemic areas [ 71. The recent cloning of HEV [ 31 led to the development of an enzyme immunoassay (EIA) to detect antibodies to recombi-

Abbreviations: HA. Enzyme immunoassay; HAV, hepatitis A virus: HCV. hepatitis C virus: HCVAb. antibody to hepatitis C virus: HEV. hepatitis E virus: HEVAb, antibody to hepatitis E virus: OW. open reading frames: SCA, sickle cell anaemia.

Correspondence: Professor S. Ramia, Department of Pathology (32) . College ofMedicine, P.O. Box 2925, Riyadh 11461, Saudi Arabia.

nant expressed HEV antigen (HEVAb) [18] and allows direct diagnosis of HEV infection.

Although the transmission of HEV is mainly enteral. some investigators [9] showed a relatively high prevalence of HEVAb in patients who received blood transfusions, implying a possible transmission of HEV through blood. In the light of our recent report [ 101 regarding the endemicity of HEV in Saudi Arabia, it was of interest, therefore, to investigate the prevalence of HEVAb in multiply trans- fused Saudi patients with sickle cell anaemia (SCA) and thalassemmia. The seroprevalence of HEV in these patients is important as high morbidity has been reported in patients with SCA and hepatitis A v i r u s (HAV) infection [ 1 1,121, another enterally transmitted hepatitis virus.

SUBJECTS AND METHODS

Patients studies

Fifty Saudi children (28 boys, 22 girls: age range 2-14 years) from the Gizan area (South), with SCA, and 28

8 1996 Blackwell Science Ltd

204 1. Al-Fawaz et al.

Saudi children (12 boys, 16 girls: age range 2-12 years) from the Riyadh area (Central province), with p- thalassemia major, who attended the haematology clinic at King Khalid University Hospital (KKUH), Riyadh, were involved in this study. Diagnosis of SCA and kthalassemia was confirmed by haemoglobin electrophoresis on cellulose acetate at pH 8.6 and cit- rate agar at pH 6.0. Adult and foetal haemoglobin were estimated by the elution method and alkaline denatu- ration, respectively, as described [12].

Healthy subjects

As the patients with SCA were from the South, the control group used was healthy children from the same area (a total of 165 children: 90 boys, 75 girls: age range 1-12 years). the control group for the tha- lassemic patients was healthy children from the Riyadh area (143 Saudi children: 80 boys, 63 girls: age range 1-14 years). The controls were investigated in another research project regarding the epidemiol- ogy of HEV infection in Saudi Arabia [ lo] . The sam- pling strategy of these children has already been described [ 1 31.

SeroZogicaZ investigations

Testing for HEV IgG was performed using the commer- cially available enzyme-linked immunosorbent assay (ELISA) (Abbott Laboratories, North Chicago, IL). In brief, two recombinant antigens (SG-3 and 8-5) derived from different open reading frames (OW) of the Burmese strain of HEV and expressed in Escherichia coli are used as solid phase antigens. The recombinant anti- gen SG- 3 is composed of 3 7 amino acids (aa) from the carboxy-terminus of ORF2, which is thought to encode

the major structural proteins of HEV [14-161. The recombinant antigen of 8-5 consists of 123 aa and rep- resents the full-length ORF3.

Antibody to HCV (HCVAb) was assayed using another commercial EIA test employing synthetic HCV peptides (UBI HCV EIA4.0; Organon Teknika, Beijing). Sera found positive for HCVAb by EIA were confirmed as true positives using a commercial line immunoassay (LiaTek; Organon Teknika).

Statistical analysis

Fisher’s exact test and t-statistic to compare propor- tions drawn from two samples were used.

RESULTS

The prevalence of HEVAb and HCVAb in multitrans- fused Saudi patients with SCA and P-thalassemia major is shown in Table 1. HEVAb among patients with SCA (18.0%) was not significantly higher than that among the control group (5.5%). Similarly, there was no sig- nificant difference in the prevalence of HEVAb between thalassemics (10.7%) and the control group (2.8%). In contrast to the situation with HEV infection, there was a significant statistical difference in the prevalence of HCVAb in patients with SCA (16.0%), P-thalassemia major (5 7.1%) and their respective control groups (1.8% and 1.4%, respectively) (Table 1).

DISCUSSION

It is well known that patients who receive multiple transfusions, such as haemophiliacs [ 141, patients on haemodialysis [15], and those with thalassemia [ 1 h] are at high risk of infection with HCV. The results of this

Table 1 Prevalence of antibodies to HEV (HEVAb) and HCV (HCVAb) in multitransfused Saudi patients with sickle cell anaemia (Gizan area) and 0-thalassemia major (Riyadh area)

Gizan area Riyadh area ~~

Sickle cell anaemia Control P-thalassemia major Control (n = 50) (n = 50) (n = 28) (n = 143) NO. (Yo) NO. (Yo) P value NO. (Yo) No. (‘+A) P value

HEVAb 9 (18.0) 9 (5.5) NS 3 (10.7) 4 (2.8) NS HCVAb 8 (16.0) 3 (1.8) 0.0078 16 (57.1) 2 (1 .4) 0.00001

NS, not significant.

0 1996 Blackwell ScienceLtd.~ournalo~ViralHepatitis. 3,203-205

HEV inpatients with SCA andPthalassaemia 205

study confirm our earlier observation [13] of the high exposure rate to HCV in two Saudi patient groups: SCA and P-thalassemia major. The exposure rate to HCV was significantly higher in patients with P-thalassemia major (57.1%) than in patients with SCA (16.0%). Frequent hospital admission and hence more blood transfusion in thalassemics may explain the difference in exposure to HCV between the two groups.

Our data show that the difference in HEV seroposi- tivity between Saudi patients with P-thalassemia major (10.7%) and their control group (2.8%), and between SCA patients (1 8.0%) and their respective control group (5.5%) is not statistically significant. This, however, does not exclude the possibility of HEV transmission via blood transfusion as the num- ber of patients in each category investigated was small and a larger number of patients should be investigated. Furthermore, the relationship between HEV seropositivity and blood transfusion should be evaluated in the light of the cumulative number of transfusions in the patients. Unfortunately this could not be satisfactorily determined in our patients as most had been transfused at more than one centre and at different occasions.

Recent studes from Saudi Arabia have shown a rela- tively high morbidity and mortality in patients with SCA and HAV infection [12,17]. Owing to the recent availability of a new test for diagnosing HEV infection, future studies are required to determine whether HEV leads to serious clinical disease in the two categories of patients studied. Sickle cell patients are susceptible to bacterial infections, especially pneumococcal sepsis. This is mainly a result of immunological dysfunction caused by poor function of the spleen [18]. Whether these patients also have poor protection against viral infections is not yet known.

The increased prevalence of HEV in multiply trans- fused patients (although not statistically significant) raises the possibility that this virus may be transmit- ted by blood products. Because of the lack of a cell cul- ture system for isolating the virus, future studies using an HEV-specific polymerase chain reaction technique may reveal whether HEV transmission through blood preparations or transfusion is possible and consequently, whether preventive measures, as with other blood-borne viruses, should be imple- mented. The results of this study need to be confirmed by extending the sample size and conducting studies in other countries.

0 1996 Blackwell Science Ltd, Iournal ojViral Hepatitis, 3,203-205

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