A serological study of cytomegalovirus and herpes simplex virus infections in Peninsular Malaysia

Bulletin of the World Health Organization, 59 (6): 909-912 (1981) A serological study of cytomegalovirus and herpes simplex virus infections in Peninsular Malaysia DORA S. K. TAN & H. STERN2 Healthy Malaysiansfrom various parts ofPeninsular Malaysia were examinedfor CF antibodies against cytomegalovirus (CMV) and herpes simplex virus (HSV) type 2. CMV antibodies were detected in 1114 out of 1556 persons (71.6%) and HSV antibodies were detected in 954 persons out of 1554 (61.4o%). The age distribution patterns were similarfor the two infections, with maximum prevalence at 5 - 14 years ofage. Prevalence was higher in women than in men. There were no significant differences among the Malay, Chinese, and Indian groups of the population with respect to CMV, 72- 781% possessing anti- bodies, but in the case of HSV, 76% of the Chinese had antibodies, compared with 57- 60% of the Malays and Indians. More than 90% of newborn infants had CMV and HSV CF antibodies, confirming the highly immune status of childbearing women in Malaysia. No CMV-specific IgM was detected in the Malaysian neonates examined but this does not exclude the possibility of congenital infection. Cytomegalovirus (CMV) and herpes simplex virus (HSV) have been recognized as causes of congenital disease, the former more so than the latter (9), but clinical cases are rarely seen in Malaysia; they are possibly missed or misdiagnosed. Cases of the acquired form are also rarely seen. The present study was undertaken to establish the prevalence of these infections in the normal popu- lation of Peninsular Malaysia. MATERIALS AND METHODS Source of sera During the period 1961 - 79, 1556 sera were collected from normal persons of various ages over the whole of Peninsular Malaysia. In addition, 264 cord-blood specimens were collected from the Chinese Maternity Hospital in Kuala Lumpur. Techniques Complement-fixing (CF) antibodies were esti- mated by the microtitration technique (14), using two units of antigen, 3 MHD5o of complement, and overnight incubation at 4 IC. All sera were inacti- vated at 56 'C for 30 min before testing. The CMV ' Senior Virus Research Officer, Virus Research Laboratory, Institute for Medical Research, Kuala Lumpur, Malaysia. 2 Professor of Virology, Department of Medical Microbiology, St George's Hospital Medical School (University of London), London, S.W.17, England. antigen was prepared from CMV-infected human embryonic lung fibroblast tissue cultures by the alkaline-glycine extraction method (3). The HSV (type 2) antigen was grown in tissue culture and supplied by the Central Public Health Laboratories, Colindale, London. The HSV CF antigen was prepared as an ultra- sonicated extract of Vero tissue culture cells infected with the Stoker strain of HSV type 1, the antigen reacts groups specifically in CF tests with both type 1 and type 2 antibodies. All sera were screened in CF tests at 1:8, and in immunofluorescence tests for IgM antibody at 1:4. IgM antibodies were determined by the indirect immunofluorescence technique, with CMV-infected fibroblasts mounted on Teflon-coated slides. Each slide was incubated with patient's serum overnight at 4 °C and then thoroughly washed and treated with anti-human-IgM conjugate (Wellcome Reagents) before examination for fluorescence by ultraviolet microscopy. Positive and negative serum controls were included in each test. Tests were regarded as positive only if immunofluorescence persisted after absorption of the test serum with staphylococcal protein A, at a titre of 32 or above. RESULTS AND DISCUSSION In many virus infections, the CF antibody response is transient, while neutralizing antibodies persist for many years. However, in the case of 4133 909 - D. S. K. TAN & H. STERN Table 1. Racial distribution of CMV and HSV CF antibodies in the Malaysian population Newborns (cord blood) Other age groupsa Total Race No. No. % No. No. % No. No. % positive examined positive positive examined positive positive examined positive CMV antibodies Malay 96 96 100.0 366 520 70.4 462 616 75.0 Indian 26 27 96.3 352 498 70.7 378 525 72.0 Chinese 110 110 100.0 396 538 73.6 506 648 78.1 Total 232 233 99.5 1114 1556 71.6 1346 1789 75.2 HSV antibodies Malay 82 96 85.4 275 519 53.0 357 615 58.0 Indian 26 27 96.3 291 498 58.4 317 525 60.4 Chinese 102 110 92.7 388 537 72.3 490 647 75.7 Total 210 233 90.1 954 1554 61.4 1164 1787 65.1 a Ages ranged from 8 days to more than 55 years. herpes viruses, including HSV and CMV, which cause lifelong latent infections, CF antibodies persist for life and therefore measurement of the titres in epidemiological surveys will give information about past infection (12). Out of 1556 Malaysians examined for CMV anti- bodies, 1114 (71.6%) were positive, and of 1554 examined for HSV antibodies 954 (61.4%) were positive (Table 1). ready spread, i.e., crowding, poor hygiene, etc. The Malaysian population obviously differs from most European and North American populations, but resembles those of Asia and Africa, in that the peak prevalence occurred at 5 - 14 years of age rather than at 25 - 35 years of age (4, 18). A slight decline in the prevalence of both antibodies occurred with age above 15 years, but was followed by a small rise after 45 years of age. Age distribution of antibodies The age distributions of the persons found to be positive for CMV antibodies and of those positive for HSV antibodies were very similar, although the prevalence of CMV antibodies was slightly high at all ages (Fig. 1). The prevalence of antibodies in chil- dren 0- 4 months old was high, 76% for CMV and 47%7o for HSV, and the persistence of these high levels in the group aged 5- 11 months, without the fall-off that might have been expected with the loss of maternal antibodies, can probably be accounted for by a high frequency of perinatal (CMV) and early postnatal (CMV and HSV) infections, originating from reactivated latent infections in the pregnant mother (8,10). CF antibodies have a tendency to fall gradually to undetectable levels after infection in early infancy and this may account for the sub- sequent dip in the curves at 1-4 years of age (15). The major rise in the prevalence of antibodies in the group aged 5- 14 years was obviously related to the extensive reservoir of infection in the child popu- lation and the existence of conditions conducive to I 00 o- so- UJ > 70 0 60 z L0- I -. I ', M.CV _HSV .4 ItE r r W1 IAGE GROUPS Fig. 1. Age distribution of CMV and HSV CF antibodies. ,2n *. * Iv. I TT__--1 910 m 114, w so I a. I 1 ..' CYTOMEGALOVIRUS AND HERPES ANTIBODIES IN MALAYSIANS I 00 90 - ~ 0~~ \ z/ = 60- 50 40 e0 30 E E AGE GROUPS Fig. 2. Distribution of CMV CF antibodiE groups. Sex distribution Prevalence of antibodies was highe 15-35 years than in men of the san CMV and HSV (Fig. 2 and 3). SimiL been reported for CMV by Luby & ' may be due to the closer contact th with children, and perhaps also to th4 latent infections during pregnancy, w maintain detectable levels of antibod ,00- 90- so- w 70 I- zri 0 'L 60 z UJ ,r 50 40 // i ! E lz E 2 vi AGE GROUPS Fig. 3. Distribution of HSV CF antibodiE groups. Race distribution Table shows that there were no significant differ- ences in the overall incidence of CMV antibodies among Malays, Indians, and Chinese (P> 0.05), and that HSV antibodies were significantly more frequent in Chinese than Malays and Indians (P< 0.001). The reason for this difference is not obvious. Incidence of antibodies in neonates \ I \I The extremely high incidence of CMV (99.507o) and * * MALE HSV (90.l1o) CF antibodies in the neonates of all FEMALE three races (Table 1) confirmed the high prevalence of immunity among childbearing women. in Malaysia (21). The fall in the prevalence of antibodies to 50-70%o at age 4 months (Fig. 2 and 3) is un- doubtedly due to loss of maternal antibodies partially counterbalanced by the acquisition of new antibodies as a result of perinatal or early postnatal infections. The presence of CMV-specific IgM was investi- .s by sex and age gated in 264 cord-blood specimens from neonates andin 96 specimens from infants 0- 4 months old. None of them were positive for IgM antibody. This does not, however, exclude the possibility of intrauterine or congenital infection in the Malaysian population. r in women aged The immunofluorescence test for CMV IgM antibodyie age, for both is relatively insensitive and commonly fails to detect ar findings have such antibody in infants with asymptomatic con- Shasby (6). This genital infection (1, 5, 7). At present, only culture of at women have the urine for virus is reliable for diagnosing congenital e reactivation of CMV infection. hich may help to The prevalence of congenital infections in the ly. highly immune populations of Alabama, USA, and of the Ivory Coast, is of the order of 1.5 - 2.5 0o (13, 16). These infections are, however, predominantly caused by reactivation of latent infection in the mother during pregnancy, and usually gives rise to perinatal infection but sometimes to true congenital infection. In the crowded populations of Asia and Africa most congenital infections are probably due to --, \ such reactivation and the situation is likely to be very ' " similar in Malaysia. Fortunately, since, in these ,>< circumstances, the fetus receives from the mother not ' only virus but also maternal antibody, the infections are usually asymptomatic and probably without residual harmful effects (16). * * MALE However, even in Alabama where, as in Malaysia, FEMALE 90% ofwomen of childbearing age possess CMV anti- bodies, primary infection does occasionally occur and, indeed, pregnant women who are seronegative ' are at particular risk of primary infection because of ,>,,>, ^ the large reservoir of infection in such populations - (19). In Alabama, primary infections accounted for about 1007o of all congenital infections (17). Primary ms by sex and age infection, unlike reactivation infection, can cause severe fetal brain damage (2, 11, 17, 20). Neverthe- Iu-. 911 912 D. S. K. TAN & H. STERN less, in Malaysia, where, also, about 1001 of women of childbearing age are seronegative for CMV anti- bodies, congenital infection is unlikely to be a signifi- cant cause of brain damage as compared with, for example, the United Kingdom, where 40 - 6007o of women of childbearing age are seronegative and most congenital infections are likely to be due to primary infection. ACKNOWLEDGEMENTS The authors wish to thank the Director of the Institute for Medical Research, Kuala Lumpur, for permission to publish this paper and Miss Yvonne Tryhorne for her technical assistance. RESUME ETUDE SEROLOGIQUE DES INFECTIONS A CYTOMEGALOVIRUS ET A VIRUS DE L'HERPES EN MALAISIE PENINSULAIRE Chez des Malais en bonne sante, originaires de diverses regions de Malaisie peninsulaire, on a recherche la presence d'anticorps fixant le complement (FC) anti-cytomegalovirus (CMV) et anti-virus de l'herpes (VH) type 2. Des anticorps anti-CMV ont e d6celes chez 1114 personnes sur 1556 (71,6%) et des anticorps anti-VH ont e trouves chez 954 sujets sur 1554 (61,4%). La distribution en fonction de l'age etait similaire pour les deux infections, avec une prevalence maximale entre 5 et 14 ans et une predominance dans le sexe feminin. Entre les groupes malais, chinois et indiens formant la population, on n'a constate aucune difference sensible sous le rapport du CMV, 72- 78% des sujets posse- dant des anticorps, mais dans le cas du VH, 76% des Chinois etaient porteurs d'anticorps, contre 57-60% des Malais et des Indiens. Plus de 90% des nouveau-nes avaient des anticorps FC anti-CMV et anti-VH, ce qui confirme le degre d'immunite eleve des femmes enceintes en Malaisie. Aucune IgM specifique du CMV n'a et decelee chez les nouveau-nes malais examines, mais cela n'exclut pas la possibilite d'une infection congenitale. REFERENCES 1. AHLFORS, K. ET AL. Acta paediatrica Scandinavica, 67: 321-326 (1978). 2. 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