Journal of Medical Virology 38195-199 (1992)
Prevalence of Herpesvirus, Human T-Lymphotropic Virus Type 1, and Treponemal Infections in Southeast Asian Refugees Dedra Buchwald, Thomas M. Hooton, and Rhoda L. Ashley Departments of Medicine (D.B., T.M.H.) and Laboratory Medicine (R.L.A.),University of Washington, Seattle, Washington Sera obtained for treponemal serology (VDRLI from 193 Southeast Asian refugees representing five ethnic groups seen i n a primary care clinic were examined for antibodies t o human T-lyrnphotropic virus type 1 (HTLV-I), human herpesvirus-6 (HHV-6), Epstein-Barr virus (EBV), and cytomegalovirus (CMV). The seroprevalence was highest for EBV (99Y0), followed in decreasing order by CMV (95%), HHV-6 (26%), and HTLV-1 (0.6%). The VDRL was positive in 15% of patients. The highest seroprevalence t o HHV-6 was noted in the Chinese (33%) and the lowest in the Laotian hilltribes, the Mien and Hmong (14%). Antibody t o HHV-6 was most prevalent among patients under 20 and those between 60 and 69 years of age. Differences were not found among ethnic groups in the seroprevalence of HTLV-1, EBV, or CMV. o 1992 Wiley-Liss,tnc.
KEY WORDS: human
herpesvirus-6, seroprevalence, seropositivity
INTRODUCTION Human T-lymphotropic virus type 1 (HTLV-1) is a recently recognized retrovirus associated with adult T cell leukemia/lymphoma and tropical spastic paraparesisiHTLV-1 associated myelopathy [Kim and Durack, 1988; Blattner et al., 19831. Although HTLV-1 is endemic primarily in areas of Japan, the Caribbean, and Africa, it is thought to occur worldwide [Kim and Durack, 1988; Blattner et al., 19831. The seroprevalence of antibodies to HTLV-1 increases with age [Blattner et al., 1983; Hinuma et al., 1982; Wang et al., 19881and sex [Wang et al., 19881. Transmission occurs transplacentally [Komuro et al., 19831, through blood transfusions [Jason et al., 19851 or by breast milk containing HTLV-l-infected lymphocytes [Kinoshita et al., 19851. Human herpesvirus-6 (HHV-6), cytomegalovirus (CMV), and Epstein-Barr virus (EBV) are ubiquitous human herpesviruses. Primary CMV and EBV infections may result in infectious mononucleosis but are more likely to occur early in life as silent infections, 6 1992 WILEY-LISS. INC.
particularly in less developed areas. Although the disease spectrum of HHV-6 is not entirely known, infection may result in a mononucleosis-like illness [Steeper et al., 19901,hepatitis [Tajiri et al., 19901,roseola infantum [Yamanishi et al., 19881or, more often, asymptomatic seroconversion in infants and children [Enders et al., 19901. All three of these herpesviruses can be isolated from saliva and all result in a latent infection from which the virus may reactivate. Little is known about the prevalence of HTLV-1 or HHV-6 in Southeast Asian populations either in the United States or in their native countries. This is due, in part, to the unavailability of testing in Southeast Asia, a culturally derived reluctance to have venipunctures performed, and the lack of accessible Southeast Asian refugee populations in the United States. The Refugee Clinic at the University of Washington is a unique primary care clinic that provides medical care to a large population of Southeast Asians in Seattle. We therefore evaluated sera from 193 refugees undergoing routine serologic screening for syphilis in a primary care clinic. Sera was tested for antibodies to HTLV-1 and HHV-6 and, as points of comparison, for antibodies to CMV and EBV.
PATIENTS AND METHODS The Refugee Clinic is located a t Harborview Medical Center, one of four hospitals affiliated with the University of Washington School of Medicine. This clinic provides medical and mental health services to a diverse population of Southeast Asian refugees from Cambodia, Laos, and Vietnam living in the greater Seattle area. The clinic has a Southeast Asian staff and 12 medical interpreters who speak 13 languages. As part of the standard evaluation, new patients undergo a complete physical examination and venipuncture for routine laboratory tests including a screening VDRL. Due to their cultural similarities and the small number of patients, two Laotian highland tribes, the Mien and Hmong, were considered as a single group. Accepted for publication April 24, 1992. Address reprint requests to Dr. Dedra Buchwald, Harborview Medical Center, 325 9th Avenue, ZA-60, Seattle, WA 98104.
Buchwald et al.
196 TABLE I. Patient Characteristics by Ethnic Group No. ( % I Mean age
Cambodian
Laotian
Mien/Hmong
81 (42)
47 (24)
22 (11)
41 17-69
41 17-77 45
53 21-79 59
Years
Range Male sex (%)
46
Chinese
Total
22 (11)
21 (11)
193
39 19-77 45
41 20-71 52
Vietnamese
42 17-79 48
TABLE 11. Seroprevalence of HTLV-1, HHV-6, CMV, EBV, and VDRL in Southeast Asian Refugees bv Ethnic G r o w Ethnic No.
group
81 Cambodian 47 Laotian MieniHmong 22 22 Vietnamese 21 Chinese 193 Total
HTLV-1
HHV-6
CMV
EBV
VDRL
80181 (99) 16180 (20) 25179 (31) 72179 (91) 0175 ( 0 ) 47147 (100) 6/46 (13) 10147 (21) 45147 (96) 1143 ( 2 ) 22122 (100) 6122 (27) 3122 (14) 21122 (95) 0121 (0) 5122 (23) 21121 (100) 21121 (100) 2/21 (10) 0121 ( 0 ) 7121 (33) 21121 (100) 21/21 (100) 0121 ( 0 ) 0118 ( 0 ) 11178 (0.6) 501191 (26) 1801190 (95) 1911192 (99) 301190 (16)
TABLE 111. Seroprevalence of HTLV-1, HHV-6, CMV, EBV, and VDRL in Southeast Asian Refugees by Age Age
No.
HTLV-1
HHV-6
CMV
EBV
70
5 83 42 34 19 10 193
015 (0) 0/79 10) 0/38 (0) 0131 ( 0 ) 1116 (6) 019 ( 0 ) 11178 (.6)
415 (80) 26/81 (32) 10/42 (23) 3/34 (8) 7119 (36) 0110 (0) 501191 (26)
5/5 (100) 78/80 (98) 38142 (90) 31/34 (91) 18/19 (95) 10/10 (100) 1801190 (95)
5/5 1100) 81/82 (99) 42142 (100) 34134 (100) 19/19 (100) 10110 (100) 1911192 (99)
Total
Sera were obtained from 193 subjects. Syphilis serologies were performed using well-established methods [U.S. Communicable Disease Center, 19691. Sera were tested anonymously for HTLV-1, HHV-6, CMV, and EBV on 178, 191, 190, and 191 patients, respectively. An enzyme-linked immunosorbent assay (ELISA) was used to test sera for antibodies to HTLV-1 (E.I. Dupont, D. Nemours, Wilmington, DE) according to the manufacturer’s instructions. Reactive specimens were tested by Western blot using antigen obtained from Hillcrest Biologicals (Cyrus, CA). HHV-6 antibodies were detected by ELISA using antigen from HHV-6 and mockinfected lymphocytes [Ashley et al., 1988al. Sera were diluted in serial twofold increments. Specimens nonreactive at the 1 : l O O dilution were considered negative. The presence of serum antibodies to CMV was determined by latex agglutination (Whittaker M.A. Bioproducts, Walkersville, MD) at a 12 dilution of sera. EBV serology was performed a t a 1:lO serum dilution by immunofluorescence for IgG antibodies to EVB capsid antigen (Meloy Laboratories, Springfield, VA).
RESULTS The demographic characteristics of the 193 study patients are shown in Table I. The group included 81 (42%) Cambodians, 47 (24%)Laotians, 22 (11%) Mien and Hmong, 22 (ll%)Vietnamese, and 21 (11%) Chi-
VDRL 015 (0) 5/82 (6) 10140 (40) 7134 (21) 8119 (42) Oil0 ( 0 ) 301190 (16)
nese. The mean age for the study population was 42 years, ranging from a mean of 39 years for the Vietnamese to a mean of 53 years for the Mien and Hmong. Forty-eight percent of the patients were male. Seroprevalences were tallied by ethnic group (Table 11).Only one patient, a Laotian female, had antibodies to HTLV-1. HHV-6 antibodies were detected in 50 (26%) of 191 patients tested. The prevalence differed among ethnic groups ranging from 149%of the Mien and Hmong to 33% of the Chinese. Overall, 22 (24%)of 92 males and 28 (28%) of 99 females had antibodies to HHV-6. In contrast, infection with CMV and EBV was virtually ubiquitous; antibodies to CMV were found in 180 (95%) of 190 patients and to EBV in 191 (99%)of 192 patients tested. The VDRL was positive in 30 (16%) of 190 patients. In 25 (83%)VDRL-positive patients the fluorescent treponemal antibody or hemagglutination test was positive. Of those with a positive VDRL, 13 (43%)were male and 17 (57%)were female. Seroprevalence data also were analyzed by subject age (Table 111). The single serum sample positive for HTLV-1 was obtained from a patient between 60 and 69 years of age. Among the 5 patients less than 20 years of age, 4 (80%) had antibodies to HHV-6. In patients age 20 years or more, the prevalence of HHV-6 antibodies ranged from 0% in the 10 individuals over 70 years to 37% in the 60-69-year-old. CMV antibodies were de-
197
Viral Seroprevalence in SE Asian Refugees TABLE IV. Seroprevalence of HTLV-1 and HHV-6 in Pacific Island and Asian Populations No. tested Location HTLV-1 Pacific Islands GuamiMicronesia 164-364
Polynesiaa
78-601
Prevalence (%a) 0.6-0.8
CL32
Reference
Brindle et al. 119881; Mora et al. 119881 Brindle et al. [19881; Hinuma et al. [19831; Asher et al. [l9881; Babona and Nurse 119881: Armstrong et al.
imoi'
Y
50-7 16
0.7-26
Brindle et al. 119881; Currie et al. 119891: Asher et al. [1988];Babona and Nurse [1988];Kazura et al. [1987]
TaiwanlChina
77-7,278
0.5-1.9
OkinawalJapan
40-343 373
20-37 0
Wang et al. [19881; Hinuma et al. 119831 Blattner et al. [1983, 19861 Hinuma et al. 119831
85 62-1,323
3.6-7 0-2.5
Melanesiab
Asia
Korea
Southeast Asia Hmong refugees Philippines
de-The et al. [19851 Ishida et al. L19881; Brindle et al. [19881; Hinuma et al. 119831
IndonesiaI Indonesian
132-158
0-45
[1989]
New Guinea
HHV-6 Japan
Hinuma et al. [19831; Asher et al. [19881; Re et al.
179-325
70-95
Yanagi et al. 119901; Hausler et al. [19901; Okuno et al.
273
42
Balachandra et al. 119891 Yadav et al. [19911
119891
Thailand Malaysia
151
58-80 (GS strain) 49-76 (229 strain)
"Includes the Solomon Islands. 'Includes Papau and Vanuatu.
Vanuatu [Hinuma e t al., 19831, Southwestern Japan [Hinuma e t al., 19821, and the Solomon Islands [Armstrong et al., 19901. In the latter study, the highest proportion of seropositive individuals were living on the most remote atoll [Armstrong et al., 19901. Others DISCUSSION also have noted higher rates of infection among isolated HTLV-1 antibody has been found in a large number populations [Ishida et al., 19881. Likewise, a high prevof Pacific Island and Asian populations using several alence was demonstrated in another geographically isomethods (Table IV). The low prevalence of HTLV-1 we lated area in Indonesian New Guinea where 45% of found in Southeast Asian refugees is comparable to subjects were seropositive [Re et al., 19891. Of greatest direct relevance to this study, two invesrates observed in several of these reports. Seroprevalences of 0-0.8% have been reported in community sur- tigations have examined the seroprevalence of HTLV-1 veys in the Philippines [Ishida et al., 19881, Micronesia in Asian immigrants and refugees. Blattner et al. and Polynesia [Brindle et al., 19881, Guam [Mora e t al., I19861 found 20% of Japanese immigrants to Hawaii 19881,Taiwan [Wang et al., 1988; Hinuma et al., 19831, from Okinawa were seropositive with low rates for imand Korea [Hinuma et al., 19831. Slightly higher rates migrants from non-endemic areas of Japan. Perhaps of 1-3% have been reported by other investigators in most comparable to our sample, de-The reported 4% of various populations of Papua [Currie et al., 19891,Asia, Hmong newcomers and 7.0% of Hmong residents in French Guyana were seropositive for HTLV-1 [de-The and the Solomon Islands [Hinuma e t al., 19831. Substantially higher HTLV-1 seroprevalences have et al., 19851. In contrast to HTLV-1, little is known about the prevbeen observed in populations residing in relatively close geographical proximity to Southeast Asia. Sero- alence of HHV-6 in Asian populations. In a study of prevalences ranging from 8-28% have been found in Southeast Asians in their homeland, Balachandra et al. the Southwestern Pacific Islands [Asher e t al., 19881, [ 19891 tested single, married, and pregnant Thai Papua [Babona and Nurse, 1988; Kazura e t al., 19871, women and found 41%, 45%, and 42%, respectively,
tected in over 90% of individuals in all age groups. The frequency of a positive VDRL increased with age until the age of 69 years, ranging from 0% in those under 20 years to 42%of those between 60 and 69 years.
Buchwald et al.
198 were seropositive for HHV-6. Although living in relatively close proximity to our patients' countries of origin, HHV-6 antibodies were substantially more common in these Thai subjects. Yadav et al. [1991] have reported evidence for two strains of HHV-6 in several diverse Malaysian populations using a n IFA assay; antibody prevalence to the 229 strain was 49-76% and 58-80% for the GS strain. In contrast, 70-96% of Japanese children and adults are seropositive for HHV-6 [Yanagi et al., 1990; Hausler et al., 1990; Okuno e t al., 1989 I. Compared to other populations the HHV-6 seroprevalence of 2610 in our group of Southeast Asian refugees is low. Antibodies to HHV-6 have been detected in over 80% of subjects from the United States as well as those from diverse countries and racial and economic backgrounds [Levy et al., 1990; Saxinger et al., 1988; Linde et al., 19881.Using the same methods reported here, we have found nearly all healthy adults of a Seattle study population were seropositive for HHV-6 [Ashley e t al., 1988133. In addition, we and others have shown that infection with HHV-6 appears to occur early in life resulting in seroprevalences of 60-8510 among infants and children [Levy et al., 1990; Linde et al., 1988; Briggs e t al., 19881. Increasing age has been associated with decreasing rates of seropositivity probably a s a result of a diminution in antibody levels [Levy e t al., 1990; Brown et al., 19881. In the current investigation, with the exception of the very elderly, we did not demonstrate this attenuation in the seroprevalence rates. The surprisingly low HHV-6 seroprevalence in our study of Southeast Asian refugees may be due to one or more of the following factors. First, the test system may be insensitive; however, this is unlikely given the high seroprevalence in American populations using this test [Ashley et al., 1988131. Second, HHV-6 may only recently have been introduced in Southeast Asia or, third, our study patients may have acquired HHV-6 infection after arriving in the United States. These possibilities seem unlikely given the high HHV-6 seroprevalence in Malaysian and Thai subjects [Balachandra e t al., 1989; Yadav et al., 19911. Finally, antigenic variants of HHV-6 strains in the countries of origin may have induced antibodies which react poorly with the prototype (African)HHV-6 strain used in our assay. Evidence for the effects of strain variation on apparent titers has been published for 229 [Yadav et al., 19911 and GS strains of HHVB [Saxinger et al., 19881. On the other hand, Suga et al. [19901 noted neutralizing antibody titers to eight strains of HHV-6 varied within a fourfold range. This observation suggested little antigenic heterogeneity but further studies were not performed. Studies employing test antigens derived from HHV-6 isolates from the appropriate country of origin are necessary to discriminate among these possibilities. Seroprevalence studies such a s this one demonstrate the marked differences in exposure to infectious agents (including different strains) among various populations. In our patients antibodies to HHV-6 were less frequent and those to CMV and EBV more frequent
than generally reported in the United States and other developed countries. Further studies are needed to determine why such dramatic differences in seroprevalences exist, whether such differences remain following immigration, and if these differences are associated with distinctive clinical manifestations among different populations.
REFERENCES Armstrong MYK, Hrdy DB, Carlson JR, Friedlaender JS (1990):Prevalence of antibodies interactive with HTLV-I antigens in selected Solomon Islands populations. American Journal of Physical Anthropology 81:465-470. Asher DM, Goudsmit J, Pomeroy KL, Garruto RM, Bakker M, Ono SG, Elliott N, Harris K, Askins H, Eldadah 2, Goldstein AD, Gajdusek DC (1988): Antibodies to HTLV-I in populations of the southwestern Pacific. Journal of Medical Virology 26:339-351. Ashley R, Abbo H, Militoni J, Corey L (1988a): Seroprevalence of human herpes virus 6 (HHV-6)by immunofluorescence and Western blot (WB). Fourteenth International Herpesvirus Workshop, Irvine, CA. Ashley R, Abbo H, Militoni J, Corey L (1988b):Seroprevalence of human herpes virus 6 (HHV-6)in four subpopulations. 28th Interscience Conference on Antimicrobial Agents and Chemotherapy, Los Angeles, p 256. Babona DV, Nurse GT (1988): HTLV-I antibodies in Papua New Guinea. Lancet 2:1148. Balachandra K, Ayuthaya PIN, Auwanit W, Jayavasu C, Okuno T, Yamanishi K, Takahashi M (1989):Prevalence of antibody to human herpesvirus 6 in women and children. Microbiology and Immunology 33515-518. Blattner WA, Blayney M, Robert-Guroff M, Sarngadharan MG, Kalyanaraman VS, Sarin PS, Jaffe ES, Gallo RC (1983):Epidemiology of human T-cell leukemiailymphoma virus. Journal of Infectious Diseases 147:406-415. Blattner WA, Nomura A, Clark JW, Ho GYF, Nakao Y, Gallo R, Robert-Guroff M (1986): Modes of transmission and evidence for viral latency from studies of human T-cell lymphotropic virus type I in Japanese migrant populations in Hawaii. Proceedings of the National Academy of Sciences USA 83:4895-4898. Briggs M, Fox J , Tedder RS (1988): Age prevalence of antibody to human herpesvirus 6. Lancet 1:105%1059. Brindle FLJ, Eglin RP, Parsons AJ, Hill AVS, Selkon J B (1988): HTLV-1. HIV-1. heuatitis B and heuatitis delta in the Pacific and Southeast Asia:' A serological survGy. Epidemiology and Infection 100:153-156. Brown N, Sumaya CV, Liu C, Ench Y, Kovacs A, Coronesi M, Kaplan MH (1988):Fall in human herpesvirus 6 seropositivity with age. Lancet 2:396. Currie B, Hinuma Y, Imai J, Cumming S, Doherty R (1989):HTLV-I antibodies in Papua New Guinea. Lancet 1:1137. de-The G, Gessain A, Gazzolo L, Najberg G, Calender A, Peti M, Brubaker G, Bensliman A, Fabry F, Strobe1 M, Robin Y , Fortune R (1985):Comparative seroepidemiologyof HTLV-I and HTLV-I11 in the French West Indies and some African countries. Cancer Research 45:S4633-54636. Enders G, Biber M, Meyer G, Helftenbein E (1990):Prevalence of antibodies to human herpesvirus 6 in different age groups, in children with exanthema subitum, other acute exanthematous childhood diseases, Kawasaki syndrome, and acute infections with other herpesviruses and HIV. Infection 18:12-15. Hausler WJ, Swack NS, Ramirez MT (1990):High prevalence of antibody to human herpesvirus-6 and decrease in titer with increase in age in Japan. Journal of Infectious Diseases 161:153. Hinuma Y, Komoda H, Chosa T, Kondo T, Kohakura M, Takenaka T, Kikuchi M, Ichimaru M, Yunoki K, Sato I, Matsuo R, Takiuchi Y, Uchino H, Hanaoka M (1982):Antibodies to adult T-cell leukemiavirus-associated antigen (ATLA) in sera from patients with ATL and controls in Japan: A nation-wide sero-epidemiologic study. International Journal of Cancer 293531-635. Hinuma Y, Chosa T, Komoda H, Mori I, Suzuki M, Tajima K, Pan I, Lee M ( 1983): Sporadic retrovirus (ATLVI-seropositiveindividuals outside Japan. Lancet 1:824-825. Ishida T, Yamamoto K, Omoto K (1988):A seroepidemiologicalsurvey
Viral Seroprevalence in SE Asian Refugees of HTLV-1 in the Philippines. International Journal of Epidemiology 17:625-628. Jason JM, McDougal JS, Cabradilla C, Kalyanaraman VS, Evatt BL (1985): Human T-cell leukemia virus (HTLV-1) p24 antibody in New York City blood product recipients. American Journal of Hematology 20:129-137. Kazura JW, Saxinger WC, Wenger J, Forsyth K, Lederman MM, Gillespie JA, Carpenter CCJ, Alpers MA (1987):Epidemiology of human T cell leukemia virus type I infection in East Sepik province, Papua New Guinea. Journal of Infectious Diseases 155:11001107. Kim JH, Durack DT (1988):Manifestations of human T-lymphotropic virus type I infection. American Journal of Medicine 84:919-928. Konoshita K, Yamanouchi K, Ikeda S, Momita S, Amagasaki T, Soda H, Ichimaru M, Moriuchi R, Katamine S, Miyamoto T, Hino S (1985): Oral infection of a common marmoset with human T-cell leukemia virus type-] (HTLV-1)by inoculating fresh human milk of HTLV-1 carrier mothers. Japanese Journal of Cancer Research (Gann)76:1147-1 153. Komuro A, Hayami M, Fujii H, Miyahara S, Hirayama M (1983): Vertical transmission of adult T-cell leukaemia virus (letter).Lancet 1:240. Levy JA, Ferro F, Greenspan D, Lennette ET (1990):Frequent isolation of HHV-6 from saliva and high seroprevalence of the virus in the population. Lancet 335:1047-1050. Linde A, Dahl H, Wahren B, Fridell E, Salahuddin Z, Biberfeld P (1988): IgG antibodies to human herpesvirus-6 in children and adults both in primary Epstein-Barr virus and cytomegalovirus infections. Journal of Virological Methods 21:117-123. Mora CA, Garruto RM, Brown P, Guiroy D, Morgan OSC, RodgersJohnson R, Ceroni M, Yanagihara R, Goldfarb LG, Gibbs CJ, Gajdusek DC (1988): Seroprevalence of antibodies to HTLV-I in patients with chronic neurological disorders other than tropical spastic paraparesis. Annals of Neurology 23:81923195. Okuno T, Takahashi K, Balachandra K, Shiraki K, Yamanishi K, Takahashi M, Baba K (1989):Seroepidemiology of human herpesvirus 6 infection in normal children and adults. Journal of Clinical Microbiology 27:651-653.
199 Re MC, Tommaseo M, Furlini G, La Placa M (1989): High prevalence of serum antibody against human T cell leukemia virus type I (HTLV-I)among the Bisman Asmat population (Indonesian New Guinea). AIDS Research and Human Retroviruses 5:551-554. Saxinger C, Polesky H, Eby N, Grufferman S, Murphy R, Tegtmeir G , Parekh V, Memon S, Hung C (1988): Antibody reactivity with HBLV (HHV-6)in U.S. populations. Journal of Virological Methods 21:199-208. Steeper TA, Horwitz CA, Ablashi DV, Salahuddin SZ, Saxinger C, Saltzman R, Schwartz B (1990):The spectrum of clinical and laboratory findings resulting from human herpesvirus-6 (HHV-6) in patients with mononucleosis-like illnesses not resulting from Epstein-Barr virus or cytomegalovirus. American Journal of Clinical Pathology 93:776783. Suga S, Yoshikawa T, Asano Y, Yazaki T, Ozaki T (1990): Neutralising antibody assay for human herpesvirus-6. Journal of Medical Virology 30:1P19. Tajiri H, Nose 0, Baba K, Okada S (1990): Human herpesvirus 6 infection with liver injury in neonatal hepatitis. Lancet 335863. U S . Communicable Disease Center (1969):Manual ofTests for Syphilis-Atlanta: Venereal Disease Program. PHS Publication No. 411. Wang CH, Chen CJ, Hu CY, You SL, Chu CT, Chou MJ, Essex M, Blattner WA, Liu CH, Yang CS (1988): Seroepidemiology of human T-cell lymphotropic virus type I infection in Taiwan. Cancer Research 485042-5044. Yadav M, Umamaheswari S, Ablashi D (1991): Antibody reactivity with two strains of human herpesvirus-6 in Malaysians. Journal of Medical Virology 33:236-239. Yamanishi K, Okuno T, Shiraki K, Takahashi M, Kondo T, Asano Y, Kurata T ( 1988):Identification of human herpesvirus-6 as a causal agent for exanthem subitum. Lancet 1:1065-1067. Yanagi K, Harada S, Ban F, Oya A, Okabe N, Tobinai K (1990):High prevalence of antibody to human herpes virus-6 and decrease in titer with increase in age in Japan. Journal of Infectious Diseases 161:153-154.
Prevalence of Herpesvirus, Human T-Lymphotropic Virus Type 1, and Treponemal Infections in Southeast Asian Refugees Dedra Buchwald, Thomas M. Hooton, and Rhoda L. Ashley Departments of Medicine (D.B., T.M.H.) and Laboratory Medicine (R.L.A.),University of Washington, Seattle, Washington Sera obtained for treponemal serology (VDRLI from 193 Southeast Asian refugees representing five ethnic groups seen i n a primary care clinic were examined for antibodies t o human T-lyrnphotropic virus type 1 (HTLV-I), human herpesvirus-6 (HHV-6), Epstein-Barr virus (EBV), and cytomegalovirus (CMV). The seroprevalence was highest for EBV (99Y0), followed in decreasing order by CMV (95%), HHV-6 (26%), and HTLV-1 (0.6%). The VDRL was positive in 15% of patients. The highest seroprevalence t o HHV-6 was noted in the Chinese (33%) and the lowest in the Laotian hilltribes, the Mien and Hmong (14%). Antibody t o HHV-6 was most prevalent among patients under 20 and those between 60 and 69 years of age. Differences were not found among ethnic groups in the seroprevalence of HTLV-1, EBV, or CMV. o 1992 Wiley-Liss,tnc.
KEY WORDS: human
herpesvirus-6, seroprevalence, seropositivity
INTRODUCTION Human T-lymphotropic virus type 1 (HTLV-1) is a recently recognized retrovirus associated with adult T cell leukemia/lymphoma and tropical spastic paraparesisiHTLV-1 associated myelopathy [Kim and Durack, 1988; Blattner et al., 19831. Although HTLV-1 is endemic primarily in areas of Japan, the Caribbean, and Africa, it is thought to occur worldwide [Kim and Durack, 1988; Blattner et al., 19831. The seroprevalence of antibodies to HTLV-1 increases with age [Blattner et al., 1983; Hinuma et al., 1982; Wang et al., 19881and sex [Wang et al., 19881. Transmission occurs transplacentally [Komuro et al., 19831, through blood transfusions [Jason et al., 19851 or by breast milk containing HTLV-l-infected lymphocytes [Kinoshita et al., 19851. Human herpesvirus-6 (HHV-6), cytomegalovirus (CMV), and Epstein-Barr virus (EBV) are ubiquitous human herpesviruses. Primary CMV and EBV infections may result in infectious mononucleosis but are more likely to occur early in life as silent infections, 6 1992 WILEY-LISS. INC.
particularly in less developed areas. Although the disease spectrum of HHV-6 is not entirely known, infection may result in a mononucleosis-like illness [Steeper et al., 19901,hepatitis [Tajiri et al., 19901,roseola infantum [Yamanishi et al., 19881or, more often, asymptomatic seroconversion in infants and children [Enders et al., 19901. All three of these herpesviruses can be isolated from saliva and all result in a latent infection from which the virus may reactivate. Little is known about the prevalence of HTLV-1 or HHV-6 in Southeast Asian populations either in the United States or in their native countries. This is due, in part, to the unavailability of testing in Southeast Asia, a culturally derived reluctance to have venipunctures performed, and the lack of accessible Southeast Asian refugee populations in the United States. The Refugee Clinic at the University of Washington is a unique primary care clinic that provides medical care to a large population of Southeast Asians in Seattle. We therefore evaluated sera from 193 refugees undergoing routine serologic screening for syphilis in a primary care clinic. Sera was tested for antibodies to HTLV-1 and HHV-6 and, as points of comparison, for antibodies to CMV and EBV.
PATIENTS AND METHODS The Refugee Clinic is located a t Harborview Medical Center, one of four hospitals affiliated with the University of Washington School of Medicine. This clinic provides medical and mental health services to a diverse population of Southeast Asian refugees from Cambodia, Laos, and Vietnam living in the greater Seattle area. The clinic has a Southeast Asian staff and 12 medical interpreters who speak 13 languages. As part of the standard evaluation, new patients undergo a complete physical examination and venipuncture for routine laboratory tests including a screening VDRL. Due to their cultural similarities and the small number of patients, two Laotian highland tribes, the Mien and Hmong, were considered as a single group. Accepted for publication April 24, 1992. Address reprint requests to Dr. Dedra Buchwald, Harborview Medical Center, 325 9th Avenue, ZA-60, Seattle, WA 98104.
Buchwald et al.
196 TABLE I. Patient Characteristics by Ethnic Group No. ( % I Mean age
Cambodian
Laotian
Mien/Hmong
81 (42)
47 (24)
22 (11)
41 17-69
41 17-77 45
53 21-79 59
Years
Range Male sex (%)
46
Chinese
Total
22 (11)
21 (11)
193
39 19-77 45
41 20-71 52
Vietnamese
42 17-79 48
TABLE 11. Seroprevalence of HTLV-1, HHV-6, CMV, EBV, and VDRL in Southeast Asian Refugees bv Ethnic G r o w Ethnic No.
group
81 Cambodian 47 Laotian MieniHmong 22 22 Vietnamese 21 Chinese 193 Total
HTLV-1
HHV-6
CMV
EBV
VDRL
80181 (99) 16180 (20) 25179 (31) 72179 (91) 0175 ( 0 ) 47147 (100) 6/46 (13) 10147 (21) 45147 (96) 1143 ( 2 ) 22122 (100) 6122 (27) 3122 (14) 21122 (95) 0121 (0) 5122 (23) 21121 (100) 21121 (100) 2/21 (10) 0121 ( 0 ) 7121 (33) 21121 (100) 21/21 (100) 0121 ( 0 ) 0118 ( 0 ) 11178 (0.6) 501191 (26) 1801190 (95) 1911192 (99) 301190 (16)
TABLE 111. Seroprevalence of HTLV-1, HHV-6, CMV, EBV, and VDRL in Southeast Asian Refugees by Age Age
No.
HTLV-1
HHV-6
CMV
EBV
70
5 83 42 34 19 10 193
015 (0) 0/79 10) 0/38 (0) 0131 ( 0 ) 1116 (6) 019 ( 0 ) 11178 (.6)
415 (80) 26/81 (32) 10/42 (23) 3/34 (8) 7119 (36) 0110 (0) 501191 (26)
5/5 (100) 78/80 (98) 38142 (90) 31/34 (91) 18/19 (95) 10/10 (100) 1801190 (95)
5/5 1100) 81/82 (99) 42142 (100) 34134 (100) 19/19 (100) 10110 (100) 1911192 (99)
Total
Sera were obtained from 193 subjects. Syphilis serologies were performed using well-established methods [U.S. Communicable Disease Center, 19691. Sera were tested anonymously for HTLV-1, HHV-6, CMV, and EBV on 178, 191, 190, and 191 patients, respectively. An enzyme-linked immunosorbent assay (ELISA) was used to test sera for antibodies to HTLV-1 (E.I. Dupont, D. Nemours, Wilmington, DE) according to the manufacturer’s instructions. Reactive specimens were tested by Western blot using antigen obtained from Hillcrest Biologicals (Cyrus, CA). HHV-6 antibodies were detected by ELISA using antigen from HHV-6 and mockinfected lymphocytes [Ashley et al., 1988al. Sera were diluted in serial twofold increments. Specimens nonreactive at the 1 : l O O dilution were considered negative. The presence of serum antibodies to CMV was determined by latex agglutination (Whittaker M.A. Bioproducts, Walkersville, MD) at a 12 dilution of sera. EBV serology was performed a t a 1:lO serum dilution by immunofluorescence for IgG antibodies to EVB capsid antigen (Meloy Laboratories, Springfield, VA).
RESULTS The demographic characteristics of the 193 study patients are shown in Table I. The group included 81 (42%) Cambodians, 47 (24%)Laotians, 22 (11%) Mien and Hmong, 22 (ll%)Vietnamese, and 21 (11%) Chi-
VDRL 015 (0) 5/82 (6) 10140 (40) 7134 (21) 8119 (42) Oil0 ( 0 ) 301190 (16)
nese. The mean age for the study population was 42 years, ranging from a mean of 39 years for the Vietnamese to a mean of 53 years for the Mien and Hmong. Forty-eight percent of the patients were male. Seroprevalences were tallied by ethnic group (Table 11).Only one patient, a Laotian female, had antibodies to HTLV-1. HHV-6 antibodies were detected in 50 (26%) of 191 patients tested. The prevalence differed among ethnic groups ranging from 149%of the Mien and Hmong to 33% of the Chinese. Overall, 22 (24%)of 92 males and 28 (28%) of 99 females had antibodies to HHV-6. In contrast, infection with CMV and EBV was virtually ubiquitous; antibodies to CMV were found in 180 (95%) of 190 patients and to EBV in 191 (99%)of 192 patients tested. The VDRL was positive in 30 (16%) of 190 patients. In 25 (83%)VDRL-positive patients the fluorescent treponemal antibody or hemagglutination test was positive. Of those with a positive VDRL, 13 (43%)were male and 17 (57%)were female. Seroprevalence data also were analyzed by subject age (Table 111). The single serum sample positive for HTLV-1 was obtained from a patient between 60 and 69 years of age. Among the 5 patients less than 20 years of age, 4 (80%) had antibodies to HHV-6. In patients age 20 years or more, the prevalence of HHV-6 antibodies ranged from 0% in the 10 individuals over 70 years to 37% in the 60-69-year-old. CMV antibodies were de-
197
Viral Seroprevalence in SE Asian Refugees TABLE IV. Seroprevalence of HTLV-1 and HHV-6 in Pacific Island and Asian Populations No. tested Location HTLV-1 Pacific Islands GuamiMicronesia 164-364
Polynesiaa
78-601
Prevalence (%a) 0.6-0.8
CL32
Reference
Brindle et al. 119881; Mora et al. 119881 Brindle et al. [19881; Hinuma et al. [19831; Asher et al. [l9881; Babona and Nurse 119881: Armstrong et al.
imoi'
Y
50-7 16
0.7-26
Brindle et al. 119881; Currie et al. 119891: Asher et al. [1988];Babona and Nurse [1988];Kazura et al. [1987]
TaiwanlChina
77-7,278
0.5-1.9
OkinawalJapan
40-343 373
20-37 0
Wang et al. [19881; Hinuma et al. 119831 Blattner et al. [1983, 19861 Hinuma et al. 119831
85 62-1,323
3.6-7 0-2.5
Melanesiab
Asia
Korea
Southeast Asia Hmong refugees Philippines
de-The et al. [19851 Ishida et al. L19881; Brindle et al. [19881; Hinuma et al. 119831
IndonesiaI Indonesian
132-158
0-45
[1989]
New Guinea
HHV-6 Japan
Hinuma et al. [19831; Asher et al. [19881; Re et al.
179-325
70-95
Yanagi et al. 119901; Hausler et al. [19901; Okuno et al.
273
42
Balachandra et al. 119891 Yadav et al. [19911
119891
Thailand Malaysia
151
58-80 (GS strain) 49-76 (229 strain)
"Includes the Solomon Islands. 'Includes Papau and Vanuatu.
Vanuatu [Hinuma e t al., 19831, Southwestern Japan [Hinuma e t al., 19821, and the Solomon Islands [Armstrong et al., 19901. In the latter study, the highest proportion of seropositive individuals were living on the most remote atoll [Armstrong et al., 19901. Others DISCUSSION also have noted higher rates of infection among isolated HTLV-1 antibody has been found in a large number populations [Ishida et al., 19881. Likewise, a high prevof Pacific Island and Asian populations using several alence was demonstrated in another geographically isomethods (Table IV). The low prevalence of HTLV-1 we lated area in Indonesian New Guinea where 45% of found in Southeast Asian refugees is comparable to subjects were seropositive [Re et al., 19891. Of greatest direct relevance to this study, two invesrates observed in several of these reports. Seroprevalences of 0-0.8% have been reported in community sur- tigations have examined the seroprevalence of HTLV-1 veys in the Philippines [Ishida et al., 19881, Micronesia in Asian immigrants and refugees. Blattner et al. and Polynesia [Brindle et al., 19881, Guam [Mora e t al., I19861 found 20% of Japanese immigrants to Hawaii 19881,Taiwan [Wang et al., 1988; Hinuma et al., 19831, from Okinawa were seropositive with low rates for imand Korea [Hinuma et al., 19831. Slightly higher rates migrants from non-endemic areas of Japan. Perhaps of 1-3% have been reported by other investigators in most comparable to our sample, de-The reported 4% of various populations of Papua [Currie et al., 19891,Asia, Hmong newcomers and 7.0% of Hmong residents in French Guyana were seropositive for HTLV-1 [de-The and the Solomon Islands [Hinuma e t al., 19831. Substantially higher HTLV-1 seroprevalences have et al., 19851. In contrast to HTLV-1, little is known about the prevbeen observed in populations residing in relatively close geographical proximity to Southeast Asia. Sero- alence of HHV-6 in Asian populations. In a study of prevalences ranging from 8-28% have been found in Southeast Asians in their homeland, Balachandra et al. the Southwestern Pacific Islands [Asher e t al., 19881, [ 19891 tested single, married, and pregnant Thai Papua [Babona and Nurse, 1988; Kazura e t al., 19871, women and found 41%, 45%, and 42%, respectively,
tected in over 90% of individuals in all age groups. The frequency of a positive VDRL increased with age until the age of 69 years, ranging from 0% in those under 20 years to 42%of those between 60 and 69 years.
Buchwald et al.
198 were seropositive for HHV-6. Although living in relatively close proximity to our patients' countries of origin, HHV-6 antibodies were substantially more common in these Thai subjects. Yadav et al. [1991] have reported evidence for two strains of HHV-6 in several diverse Malaysian populations using a n IFA assay; antibody prevalence to the 229 strain was 49-76% and 58-80% for the GS strain. In contrast, 70-96% of Japanese children and adults are seropositive for HHV-6 [Yanagi et al., 1990; Hausler et al., 1990; Okuno e t al., 1989 I. Compared to other populations the HHV-6 seroprevalence of 2610 in our group of Southeast Asian refugees is low. Antibodies to HHV-6 have been detected in over 80% of subjects from the United States as well as those from diverse countries and racial and economic backgrounds [Levy et al., 1990; Saxinger et al., 1988; Linde et al., 19881.Using the same methods reported here, we have found nearly all healthy adults of a Seattle study population were seropositive for HHV-6 [Ashley e t al., 1988133. In addition, we and others have shown that infection with HHV-6 appears to occur early in life resulting in seroprevalences of 60-8510 among infants and children [Levy et al., 1990; Linde et al., 1988; Briggs e t al., 19881. Increasing age has been associated with decreasing rates of seropositivity probably a s a result of a diminution in antibody levels [Levy e t al., 1990; Brown et al., 19881. In the current investigation, with the exception of the very elderly, we did not demonstrate this attenuation in the seroprevalence rates. The surprisingly low HHV-6 seroprevalence in our study of Southeast Asian refugees may be due to one or more of the following factors. First, the test system may be insensitive; however, this is unlikely given the high seroprevalence in American populations using this test [Ashley et al., 1988131. Second, HHV-6 may only recently have been introduced in Southeast Asia or, third, our study patients may have acquired HHV-6 infection after arriving in the United States. These possibilities seem unlikely given the high HHV-6 seroprevalence in Malaysian and Thai subjects [Balachandra e t al., 1989; Yadav et al., 19911. Finally, antigenic variants of HHV-6 strains in the countries of origin may have induced antibodies which react poorly with the prototype (African)HHV-6 strain used in our assay. Evidence for the effects of strain variation on apparent titers has been published for 229 [Yadav et al., 19911 and GS strains of HHVB [Saxinger et al., 19881. On the other hand, Suga et al. [19901 noted neutralizing antibody titers to eight strains of HHV-6 varied within a fourfold range. This observation suggested little antigenic heterogeneity but further studies were not performed. Studies employing test antigens derived from HHV-6 isolates from the appropriate country of origin are necessary to discriminate among these possibilities. Seroprevalence studies such a s this one demonstrate the marked differences in exposure to infectious agents (including different strains) among various populations. In our patients antibodies to HHV-6 were less frequent and those to CMV and EBV more frequent
than generally reported in the United States and other developed countries. Further studies are needed to determine why such dramatic differences in seroprevalences exist, whether such differences remain following immigration, and if these differences are associated with distinctive clinical manifestations among different populations.
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