Prevalence of Antibodies to Herpesviruses in Central Saudi Arabia

Prevalence of Antibodies to Herpesviruses in Central Saudi Arabia Talal M. F. Bakir, PhD* * Assistant Professor and Consultant Virologist, Department of Pathology (32), College of Medicine and King Khalid University Hospital, King Saud University, P.O. Box 2925, Riyadh 11461, Saudi Arabia Date of Acceptance: 5 October 1986 ABSTRACT Sera from a total of 770 individuals (488 adults, 282 children) living mainly in the central area of Saudi Arabia were serologically investigated for antibodies to the herpesvirus family, which is known to cause medically important diseases. The herpesviruses include: cytomegalovirus (CMV), Epstein-Barr virus (EBV), varicella zoster virus (VZV) and herpes simplex viruses (HSV-1 and HSV-2). Using the indirect immunofluorescent test, the overall prevalence of antibodies in children was 75.5% for CMV, 61% for EBV, 66.6% for VZV, 55.3% for HSV-1, and 0% for HSV-2. For adults, the overall prevalence of antibodies was 88.5% for CMV, 85.2% for EBV, 88.9% for VZV, 89.5% for HSV-1 and 3% for HSV-2. This indicates a high incidence of infection for all these herpesviruses apart from HSV-2, as well as early acquisition of infection in childhood. Males and females appeared to be almost equally susceptible to infection by the entire family of herpesviruses. TMF Bakir, Prevalence of Antibodies to Herpesviruses in Central Saudi Arabia. 1987; 7(3): 196-201 MeSH KEYWORDS: Herpesvirus infections - prevention and control - Saudi Arabia Introduction THE HERPESVIRUS family consists of medically important viruses which include cytomegalovirus (CMV), Epstein-Barr virus (EBV), varicella zoster virus (VZV), and herpes simplex viruses (HS V-1 and HSV-2). All of these exhibit a characteristic feature of causing latent infections persisting perhaps during the lifetime of the host. 1 Infection by all these herpesviruses is known to occur in childhood, through close contact and low standards of hygiene, associated often with poor economic conditions. 2,5

A wide range of infections is caused by this group of viruses. CMV causes manifestations varying from mild (e.g., mononucleosis-like syndrome) to moderate (e.g., hepatitis) to severe (e.g., congenital infections). 6,8 EBV has been implicated in infectious mononucleosis, 9 Burkitt's lymphoma and nasopharyngeal carcinoma. 10 VZV can be a serious illness in children with primary immunodeficiency and may lead to severe progressive chicken pox. 11 HSV-1 causes oral infections which can be mild or severe. In adolescents, vesicular pharyngitis and tonsillitis are common, as is gingivostomatitis in younger children, 1,12 but severe encephalitis with high mortality and morbidity is the most serious complication. 13 HSV-2 usually causes genital infections and, when sexually transmitted, carries the risk of neonatal infection through the infected birth canal at delivery. 14 Seroepidemiological reports on the prevalence of specific antibodies to the entire family of human herpesviruses in the Middle East are scarce, and little is known about their epidemiology in Saudi Arabia. The extensive area and variability of socioeconomic conditions of life in urban and rural Saudi Arabia may possibly have a major role in epidemiology and prevalence of antibodies against the entire herpesvirus family. Several serological techniques are currently in use for the detection of rising titer of specific antibodies to these viruses. In this study, using the indirect immunofluorescent antibody (IFA) test, we have investigated the prevalence of specific IgG antibodies for CMV, EBV, VZV, HSV-1 and HSV-2 in the central region of Saudi Arabia, in order to determine the age of primary infection. The indirect IFA has been shown to be a useful screening procedure, as well as being rapid, easy to perform, and more sensitive than the complement fixation test (CFT). 15 Materials and Methods Sera Samples of 770 sera from various age groups were obtained, mainly from the central region of Saudi Arabia (Riyadh), though some sera were from people coming from the Eastern Province (Dammam) and the Southern Province (Abha, Najran). The sera from adults were from healthy male blood donors and pregnant women. Children's sera were obtained from asymptomatic patients. IFA Tests Commercial test kits were obtained and the following tests carried out according to the instructions of the manufacturers (Virgo Reagents, Electronucleonics, Inc., Columbia, Maryland, USA). CMV. The IFA test for CMV-IgG antibody was used. Screening was done at 1:16 dilution. Sera which gave positive fluorescence at that dilution were considered to be positive for CMV-IgG antibodies. EBV. The EBV viral capsid antigen (EBV-VCA) IFA test was used for the detection of antibody to EBV. Screening was carried out at 1:10 dilution, and positivity was taken to be indicative of immunity to infectious mononucleosis. VZV. The IFA test for VZV-IgG antibody was used. Screening for sera was carried out at 1:8 dilution. Sera which gave positive fluorescence at that dilution were considered to be positive for VZV-IgG antibodies. HSV-1 and HSV-2. The IFA test for HSV-IgG antibody was used. This consisted of mammalian cells infected with the prototype type 1 or the prototype type 2. Sera were screened at 1:10 dilution. HSV-1 conjugate stain infected cells with an intensity of 3+ to 4+ whereas HSV-2 infected cells stained with an intensity of 2+ or less. Cells infected with HSV-1 showed strong perinuclear fluorescence compared to homogenous (diffuse) staining of the entire HSV-2 infected cells. Sera which gave positive fluorescence at 1:10 with HSV-1 but showed negative fluorescence with HSV-2 were considered to be positive for HSV-1, and sera with positive fluorescence at 1:10 with HSV-2, but no fluorescence with HSV-1 were taken to be positive for HSV-2. Results A total of 770 sera from different age groups were tested for antibodies to all human herpesviruses: CMV, EBV, VZV, HSV-1, and HSV-2. A marked variation according to age, but not sex, was obtained. The highest overall antibody prevalence in children was due to CMV (75.5%) followed by VZV (66.6%), EBV (61%), HSV-1 (55.3%) and HSV-2 (0%). In adults, the highest overall antibody prevalence was due to HSV-1 (89.5%), followed by VZV (88.9%), CMV (88.5%), EBV (85.2%) and HSV-2 (3%). CMV antibodies. Table 1 shows the results of CMV-IgG determinations carried out in the various age groups. Infection with CMV is acquired early in life, in that 73.3% of the male children and 72.7% of female children had CMV-IgG antibodies by the age of 5 years, after which there was a gradual increase in the CMV antibodies with increasing age, reaching a peak of 87.5% in males 16-25 years old.

EBV antibodies. The results of EBV-VCA-IgG are shown in Table 1. Fifty percent of male children and 45.4% of female children up to 5 years old had IgG antibodies against EBV. Then there was a gradual increase, and by the age of 9 years, 63.1% of male and 65% of female children had EBV-IgG antibodies. The age group of 10-15 years showed a higher percentage of antibodies (males 75%; females 79.5%), increasing to 88.2% in males of 26-35 years. In the older age groups, the number with antibodies remained without significant changes in both males and females. VZV antibodies. Table 1 shows the results concerning VZV antibodies. There was a gradual increase in IgG antibody against VZV. By the age of 5 years, 46.6% of male children had VZV antibodies. By age 6-9 years, 70% of female children and 73.6% of male children had VZV antibodies. There was an increase of antibodies in the 10-15 year age groups in both sexes. The peak of 90.8% in females and 91.9% in males was reached in the age group 26-35 years. Above 35 years of age, the percentage of people of both sexes with antibodies was maintained without significant changes. Table 1. Prevalence of IF-IgG antibodies to CMV, EBV, VZV, HSV-1 and HSV-2. Age (yrs) Male CMV EBV VZV HSV-1 HSV-2 No. Ab+ % Ab+ % Ab+ % Ab + % Ab+ % <1 26 17 65.3 11 42.3 12 46.1 8 40.7 0 0 1-5 30 22 73.3 15 50.0 14 46.6 12 40.0 0 0 6-9 38 30 78.9 24 63.1 28 73.6 22 57.8 0 0 10-15 48 " 39 81.2 36 75.0 41 85.4 36 75.0 0 0 Female <1 23 14 60.8 10 43.4 10 43.4 6 26.0 0 0 1-5 33 24 72.7 15 45.4 16 48.4 14 42.4 0 0 6-9 40 29 72.5 26 65.0 28 70.0 24 60.0 0 0 10-15 44 38 86.3 35 79.5 39 88.6 34 77.2 0 0 Total children 282 213 75.5 172 61.0 188 66.6 156 55.3 0 0 Male 16-25 72 63 87.5 63 87.5 64 88.8 62 86.1 3 4.1 26-35 136 119 87.5 120 88.2 125 91.9 122 89.7 8 5.8 >35 56 48 85.7 47 83.9 49 87.5 52 92.8 3 5.3 Female 16-25 61 53 86.8 50 81.9 51 83.6 51 83.6 0 0 26-35 120 111 92.5 101 84.1 109 90.8 110 91.6 1 0.83 >35 43 38 88.3 35 81.3 36 83.7 40 93.0 0 0 Total adults 488 432 88.5 416 85.2 434 88.9 437 89.5 15 3.0 HSV-1 and HSV-2 antibodies. The prevalence of HSV-1-IgG and HSV-2-IgG antibodies is shown in Table 1. HSV-1 antibodies have a prevalence of 40% and 42.4%, respectively, in 1-5-year-old males and females. Then there is a gradual increase in both males and females, reaching a peak of 93% in females and 92.8% in males over 35 years of age. This gradual increase of HSV-1 antibodies is similar to the prevalence of the rest of the herpesvirus antibodies. However, HSV-2 did not appear in childhood (<1-15 years). In early adulthood (16-25 years) a prevalence of 4.1% of HSV-2 antibodies was seen in males. This increased to 5.8% by ages 26-35 in males. In the age group >35 years, 5.3% of the males had HSV-2 antibodies. Females showed only one positive HSV-2 titer among 364 sera tested, in a woman in the 26-35 age group. It was noted from the results described in Table 1 that the Fischer exact tests were significantly different (P < 0.05) for CMV and HSV-1 antibodies in males and females in the age group <1 year. Other significant differences were noticed for acquiring antibodies to CMV on one hand and to EBV, VZV and HSV-1 on the other hand in males and females, age group 1-5 years.

Discussion This is the first broad-based investigation to determine antibodies to the entire family of herpesviruses among different sexes and age groups in Saudi Arabia, using a single sensitive and specific test. Data obtained in this study suggest that there is an early acquisition of CMV, EBV, VZV and HSV-1 infections. The risk of infection to children in age groups <1 year up to 5 years old might vary with different members of the herpesvirus family. As seen in Table 1, there is a clear difference between the early acquisition of antibodies to CMV and antibodies to HSV-1, EBV and VZV. This information suggests that, although all herpesviruses investigated in this study can be acquired relatively early in life, CMV can be acquired very early. This confirms that close contact, and possibly saliva and urine contamination, in these very young children are major causes of infection. 16 Results of CMV, HSV-1 and HSV-2 are similar to our previous study, 17 where it was found that 88% of pregnant women had CMV antibodies, 92% had HSV-1 antibodies, and 6.3% had HSV-2 antibodies. CMV. These results show that CMV infection is endemic in Saudi Arabia and that the prevalence of CMV antibodies is high. In early childhood, between the ages 1 and 5 years, it is 73%. In Saudis over 35 years old, it is 86-88%. The endemicity of CMV in Saudi Arabia, as shown in Table 1, is quite similar to other Middle Eastern and African countries. 18,19 Antibodies to CMV in infants under one year of age, as with the other herpesviruses, are probably transmitted by maternal antibodies. In this study, all of the adult population, including the pregnant women, had almost equally high rates of antibodies against CMV, and also against HSV-1, EBV and VZV. Thus, passive transplacental immunity might be transferred at the same rate for all these herpesviruses. No sex differences were noted for the different members of the herpesvirus family. Similar findings have been reported by others in different populations. 15,16,18 Despite the variability in social status among the 770 people studied, the close family contact that is part of the social structure in Saudi Arabia plays a major role in the transmission in the community of CMV and the rest of the herpesviruses. The early exposure to CMV infection in this population could explain the low incidence of congenital infection due to CMV, compared to developed countries. 16 The early acquisition of CMV antibodies in children in Saudi Arabia and high prevalence of these antibodies in adulthood agrees with reports from Europe, the United States and other parts of the world. 18,22 However, a few reports from the United Kingdom, United States and Switzerland 16,19 indicated much lower prevalence of CMV antibodies in children. One factor contributing to this low prevalence of CMV antibodies could be the low sensitivity to the CFT used in these studies. EBV. The early acquisition of EBV infection is demonstrated in Table 1. Our data show that, in the case of children, the overall incidence of antibodies to EBV was 61%, and this level increased to 85.2% in adults. These results are in agreement with previous reports. 23,25 The levels of EBV-IgG antibodies in Saudi Arabia are lower than those detected in India recently, 26 which may relate to the low standard of hygiene and endemicity of EBV in that part of the world. In Uganda, Kafuko, et al, 27 found that 90% of children had EBV antibodies by the age of 3 years, dropping to <75% by age 15. This drop of antibodies was explained by the endemicity of EBV in the young children in past years. Our results, like those of Isager, et al, 24 suggest that intense exposure to EBV plays a major role in spreading the virus in the population. On the other hand, previous reports 28 showed that delay or failure of EBV infection occurred in spite of great exposure to the virus. So, intense exposure was not the only reason for EBV infection, but perhaps additional unknown factors play a role. Neiderman, et al, 29 found that prevalence of EBV antibodies depends on age and geographic area. At Yale University in the United States, 26% of students 17-18 years of age had EBV antibodies, while at the University of the Philippines, 75% of 17-22 year olds were positive for EBV antibodies, and 87% of 18-20-year-old military recruits in Colombia had EBV antibodies. VZV. Among both male and female children there were no significant differences observed in the acquisition of VZV antibodies. Both sexes initially registered 46.1% and 43.4% positive in males and females, respectively, at <1 year of age. In adults the level of VZV antibodies remained unchanged in both sexes. From Table 1 it is noted that VZV antibodies are present in up to 88.6% of older children, indicating the early acquisition of this virus in our community. Our results agree with most studies which show that most adults will have antibodies to VZV and are immune to infection. 15,30,31 In a seroepidemiology study of VZV antibodies in women and their children in America by Gershon, et al, 30 it was found that 84% of foreign-born women and 95% of American women of childbearing age had antibodies to VZV. This suggests a higher susceptibility rate to VZV in young adults from tropical countries (e.g., Puerto Rico, Cuba) than in women born in America. Other reports also indicate that children from tropical and subtropical countries have a lower incidence of chicken pox than in temperate countries. 32 HSV-1 and HSV-2. It is clear that early acquisition of HSV-1 antibodies is significantly less than CMV in children of <l-5 years old. Only 40% of male children in the 1-5-year-old group had HSV-1 antibodies. In contrast, 73.3% of the same children had antibodies to CMV. Very similar data were obtained in other countries. 18,24 These

are in contrast to other reports 20 from investigators who used CFT for detecting HSV antibodies and found a much lower prevalence of HSV-1 antibodies in children 5 months to 16 years of age. These differences could be due to the low sensitivity of CFT. Porter, et al, 25 found a lower prevalence of HSV-1 antibody in higher than in lower socioeconomic groups. This was noticed mainly in the age group of 8-14 years, in which 40% of a high socioeconomic group had HSV-1 antibodies compared with 80% in the lower socioeconomic group. Glezen, et al, 33 found only 30% of university students had antibodies against HSV-1 and only 10% of people with no antibodies to HSV-1 will acquire antibodies each year. They suggested that most HSV-1 infections will be acquired after young adulthood by close and intimate contact. The wide spread of HSV-1 in native people in some countries, summarized by Gerber and Rosenblum, 34 indicated a very high prevalence of HSV-1 antibodies in isolated communities. HSV-2 antibodies were not detected in any children. In males in the age group 16-35 years there was a low prevalence rate of HSV-2 antibodies (4.1 to 5.8%) and in females only one case was detected in the age group 26-35 years. Our results in adult males were quite similar to Corey and Holmes, 35 who found the frequency of genital herpes simplex infections in patients attending sexually-transmitted disease clinics in Washington, USA, is 5.1%. The mode of transmission of HSV-2 is different than for HSV-1. The former usually is venereally transmitted. 14,35 On the other hand, there is cross reaction between HSV-1 and HSV-2 antibodies as they share common antigens. 36 Accordingly, detection of antibodies to HSV-2 may not be diagnostic for type 2 infection, unless the serum shows no positivity for HSV-1. References 1. Fenner FJ, White DO. Herpetoviridae. In: Medical virology. 2nd ed. New York: Academic Press, 1976:297-319. 2. Li F, Hanshaw JB. Cytomegalovirus infection among migrant children. Amer J Epidem 1967;86:137-41. 3. Scott TF. Epidemiology of herpetic infections. Amer J Ophthal 1957;43:134-47. 4. Hyams PJ, Vernon S, Eckert D. Susceptibility to varicella virus of certain adults in the southeastern United States. Am J Infec Control 1984;12(l):6-9. 5. Ginsburg CM, Henle G, Henle W. An outbreak of infectious mononucleosis among the personnel of an outpatient clinic. Am J Epidemiol 1976;104(5):571-5. 6. Klemola E, Kaariainen L. Cytomegalovirus as a possible cause of a disease resembling infectious mononucleosis. Br Med J 1965;5470:1099-1102. 7. Toghill PJ, Bailey ME, Williams R, et al. Cytomegalovirus hepatitis in the adult. Lancet 1967;1:1351-4. 8. Lang DJ, Noren B. Cytomegaloviremia following congenital infection. J Pediatr 1968;73(6):812-9. 9. Nikoskelainen J, Hanninen P. Antibody response to Epstein-Barr virus in infectious mononucleosis. Infect Immun 1975;11(1):42-51. 10. Epstein MA, Achong BG. 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The incidence of infection with cytomegalovirus in a normal population: a serologic study in greater London. J Hyg (Camb) 1965;63:79-87. 17. Hossain A, Bakir TM, Ramia S. Immune status to congenital infections to TORCH agents in pregnant Saudi women. J Trop Pediatr 1986;32(2):83-6. 18. Al-Nakib W. A modified passive-hemagglutination technique for the detection of cytomegalovirus and herpes simplex virus antibodies: application in virus-specific IgM diagnosis. J Med Virol 1980;5(4):287-93. 19. Krech U, Jung M, Jung F. Cytomegalovirus infections of man. Basel: S Karger, 1971:27-28. 20. Baron J, Youngblood L, Siewers MF, et al. The incidence of cytomegalovirus, herpes simplex, rubella and toxoplasma antibodies in microcephalic, mentally retarded and normocephalic children. Pediatrics 1969;44:932-9. 21. Haneberg B, Bertnes E, Haukenes G. Antibodies to cytomegalovirus among personnel at a children's hospital. Acta Pediatr Scand 1980;69(3):407-9. 22. Mindel A, Sutherland S. Antibodies to cytomegalovirus in homosexual and heterosexual men attending an STD clinic. Br J Vener Dis 1984;60(3): 189-92. 23. Henle G, Henle W. Immunofluorescence, interference, and complement fixation technics in the detection of the herpes-type virus in Burkitt tumor cell lines. Cancer Res 1967;27:2442-6. 24. Isager H, Cohn J, Hesse J, Vestergaard BF. Antibodies to herpes simplex and Epstein-Barr viruses in Faroese children:

association with sibship, size, height and age. Acta Pathol Microbiol Immunol Scand (B) 1983;91(5):339-41. 25. Porter DD, Wimberly I, Benyesh-Melnick M. Prevalence of antibodies to EB virus and other herpesviruses. JAMA 1969;208:1675-9. 26. Venkitaraman AR, Lenoir GM, John TJ. The seroepidemiology of infection due to Epstein-Barr virus in southern India. J Med Virol 1985;15(1):11-16. 27. Kafuko GW, Henderson BE, Kirya BG, et al. Epstein-Barr virus antibody levels in children from the West Nile District of Uganda. Lancet 1972;1:706-9. 28. Chang RS. What is the determinant for susceptibility to infectious mononucleosis? N Engl J Med 1975; 292(54): 1298. 29. Niederman JC, Evans AS, Subrahmanyan L, McCollum RW. Prevalence, incidence and persistence of EB virus antibody in young adults. N Eng J Med 1970;282:361-5. 30. Gershon AA, Raker R, Steinberg S, et al. Antibody to varicella-zoster virus in parturient women and their offspring during the first year of life. Pediatrics 1976, 58(5):692-6. 31. Ross AH. Modification of chicken pox in family contacts by administration of gamma globulin. N Engl J Med 1962;267:369-76. 32. Maretic Z, Cooray MP. Comparisons between chicken pox in a tropical and a European country. J Trop Med Hyg 1963;66:311-5. 33. Glezen WP, Fernald GW, Lohr JA. Acute respiratory disease of university students with special reference to the etiologic role of Herpesvirus hominis. Am J Epidemiol 1975;101(2):111-21. 34. Gerber P, Rosenblum EN. The incidence of complement-fixing antibodies to herpes simplex and herpes-like viruses in man and rhesus monkeys. Proc Soc Exp Biol Med 1968;128:541-6. 35. Corey L, Holmes KK. Genital herpes simplex virus infections: current concepts in diagnosis, therapy and prevention. Ann Intern Med 1983; 98(6):973-83. 36. Geder L, Skinner GRS. Differentiation between type 1 and type 2 strains of herpes simplex virus by an indirect immunofluorescent technique. J Gen Virol 1971;12:179-82.