RISK FACTORS FOR NIPAH VIRUS TRANSMISSION RISK FACTORS FOR NIPAH VIRUS TRANSMISSION, PORT DICKSON, NEGERI SEMBILAN, MALAYSIA: RESULTS FROM A HOSPITAL-BASED CASE-CONTROL STUDY NM Amal1, MS Lye1, TG Ksiazek2, PD Kitsutani2, KS Hanjeet1, MA Kamaluddin1, Flora Ong3, Sutha Devi4, PC Stockton2, O Ghazali5, R Zainab5 and MA Taha6 1Division of Epidemiology Research, Institute for Medical Research, Kuala Lumpur, Malaysia; 2Special Pathogens Task Force Head Quarters, Centers for Disease Control and Prevention (CDC), Atlanta, USA; 3State Health Department, Sarawak, Malaysia; 4Hospital Port Dickson, Negeri Sembilan, Malaysia; 5State Health Department, Negeri Sembilan, Malaysia; 6Disease Control, Ministry of Health, Malaysia Abstract. A hospital-based case-control study of viral encephalitis was carried out at Port Dickson Hospital, in the state of Negeri Sembilan, Malaysia. Between March and May 1999, 69 clinically diagnosed viral encephalitis cases and 31 controls were interviewed. Job histories on pig farming activities were assessed by a group of epidemiologists and veterinary surgeons. Results show that among clinical cases of viral encephalitis, 52 (75.4%) cases were diagnosed to have Nipah virus infection based on positive serology for antibodies to the cross-reacting Hendra virus antigen. The Nipah virus encephalitis was significantly associated with a history of working in pig farms (p < 0.001, OR = 196.0, 95% CI = 20.4 - 4741.6), history of contact with animals (p < 0.001, OR = 38.3, 95% CI = 8.2 - 209.0) and with history of direct contact with pigs (p = 0.002, OR = 34.4, 95% CI = 2.6 - 1,024.4). The Nipah virus infection was also significantly associated with history of feeding/cleaning pigs (p < 0.001, OR = 102, 95% CI = 11.9 - 2,271.5). These results provide evidence that involvement in pig farming activities is significantly associated with the risk of getting Nipah virus infection. They are potential risk factors for Nipah virus transmission in the major pig-producing area of Bukit Pelandok, Port Dickson Negeri Sembilan. INTRODUCTION Laboratory investigations of cases of encephalitis suggested of a previously unknown virus that rep- licated in pigs and readily spread to humans. The An outbreak of viral encephalitis, which re- virus was initially isolated at the University Hos- sulted in over 100 human fatalities, began in late pital in Kuala Lumpur in early March 1999. The February and peaked around the middle of March virus was later identified as a new paramyxovirus 1999 in Bukit Pelandok, Port Dickson district, closely related to the Hendra virus (formerly called Negeri Sembilan state in Peninsular Malaysia. Bukit equine morbillivirus) at the Centers for Disease Pelandok is one of the largest pig-producing areas Control and Prevention (CDC) in Atlanta, Georgia, in the country. The outbreak primarily involved USA. The Malaysian virus was subsequently named male adults with histories of direct contact with Nipah virus, named after the village Sungai Nipah pigs. Their family members who also lived but did (Nipah river) in Negeri Sembilan state, where a not work on the pig farms were not affected (Enserink, pig farm worker from which the first viral isolate 1999). Early epidemiologic investigations suggest came from had died. that the new virus is easily transmitted to humans, and spreads rapidly among pigs. By end of April Hendra virus was first recognized in 1994 1999 most pig farms in Bukit Pelandok area were after an outbreak of respiratory disease among affected. It also seemed that the new virus only horses and humans in Australia (Murray et al, affects those who had been in close contact or 1995a,b; Selvey et al, 1995). To date, two out- directly involved in pig farming activities (CDC, breaks of Hendra virus have been reported, with 1999a,b; Enserink, 1999). three human cases and two deaths (Hooper et al, 1996; Murray et al, 1995a,b; O’Sullivan et al, Correspondence: Dr Munn-Sann Lye, Institute for Medi- cal Research, Jalan Pahang, 50588 Kuala Lumpur, 1997; Rogers et al, 1996; Selvey et al, 1995). Malaysia. Hendra virus is believed to spread through direct Tel: (603) 4402302; Fax: (603) 2920675; E-mail: lyems@ contact with body fluids of infected horses and has imr.gov.my been isolated from both saliva and urine of horses Vol 31 No. 2 June 2000 301 SOUTHEAST ASIAN J TROP MED PUBLIC HEALTH (McCormack et al, 1999; Williamson et al, 1998). Data collection This study identified potential risk factors for Nipah After obtaining consent, a standardized ques- virus transmission in the district of Port Dickson, tionnaire was administered to study subjects by Negeri Sembilan state. trained members of the investigation team between 1 March and 15 May 1999. An adult family mem- ber was interviewed for deceased cases and for MATERIALS AND METHODS cases who could not be personally interviewed due to the severity of their illness. Information was Study population obtained about household, demographics and so- cioeconomic characteristics, illness, exposure to The peak of the Nipah viral disease outbreak pigs and other animals, characteristics of the farm, occurred during the middle of March and contin- and (Japanese encephalitis) vaccination history. ued through May 1999 in the state of Negeri Sembilan. According to the Ministry of Health, Collection of specimens Malaysia, most of the cases were reported from the Port Dickson district. Cases and controls for A single specimen of cerebrospinal fluid (CSF) this study were chosen from this district. Most of from case-patients and 10 ml of venous blood from the population were of Chinese ethnicity and in- cases and controls were obtained. These specimens volved in pig farming activities, particularly from were tested for presence of Nipah virus IgM and/ the highly concentrated pig farming area of Bukit or IgG antibodies. All specimens were tested at Pelandok, Negeri Sembilan state. the special National Task Force Laboratory oper- ated by the CDC at the University Hospital, Kuala Selection of cases Lumpur during the outbreak. Specimens were tested for IgM antibodies by using an IgM-capture an- Cases were defined as persons with serologi- tibody enzyme immunoassay (EIA) and for IgG cal evidence of Nipah virus infection. Participants antibodies by using an indirect EIA. Preliminary were recruited based on their residential addresses nucleotide sequence information indicated that the of Port Dickson, Negeri Sembilan, and who were Nipah virus was closely related to Hendra virus, hospitalized with a clinical diagnosis of viral en- cross-reacting in a Hendra antigen ELISA (CDC, cephalitis that were reported to the Ministry of 1999a). Therefore, in this study, Hendra virus an- Health, Malaysia, during the period from March tigens, which cross react with Nipah virus anti- through May 1999. Viral encephalitis patients who bodies, were used in the assays. had been discharged from the hospital and could be located were recruited through home visits. Data management and analysis Eligible encephalitis candidates whose serum speci- men tested positive for Nipah virus antibody and Questionnaire and serological data were en- who consented to participate in the study were tered into databases. EPI-info version 6.04b (Cen- included as cases. Clinical encephalitis patients ters for Disease Control and Prevention, Atlanta, whose specimens tested negative for Nipah virus George, USA) was used for analysis. We calcu- antibody were excluded from the study. lated odds ratio (OR), 95% confidence intervals (CI) and p-values by chi-square with Yates’correction Selection of controls or two-tailed Fisher’s exact test, when appropriate. Controls were patients of non-Muslim faith with residential addresses in Port Dickson and admitted to Port Dickson Hospital, Negeri Sembilan RESULTS during the month of March through May 1999. Muslims were excluded because their religious A total of 69 cases of viral encephalitis were beliefs and practices prohibit them from direct identified. Of these, 52 (75.4 %) were serologi- contact with pigs or consuming pork. Patients cally positive for Nipah virus. A total of 16 cases with clinical diagnoses of encephalitis, meningitis, were excluded from the study either because their pneumonia, and viral fever other than dengue, laboratory results were negative for Nipah virus measles and chickenpox were excluded. Patients (11 cases, 15.9 %) or their laboratory results were admitted to the obstetrics and gynecology wards not available (six cases, 8.7 %) (Table 1). All 31 were also excluded. controls were negative for Nipah virus infection. 302 Vol 31 No. 2 June 2000 RISK FACTORS FOR NIPAH VIRUS TRANSMISSION Table 1 Number of viral encephalitis cases included/excluded in the study of Nipah virus infection, Port Dickson, Negeri Sembilan, Malaysia, March-May 1999. Reasons for exclusion No. (%) Serology negative (IgM/IgG) for Nipah virus 11 15.9 Serology results were not available (IgM/IgG) 6 8.7 Serology positive (IgM and /or IgG) for Nipah virus 52 75.4 Total 69 100 Table 2 Age, sex, and race distribution of Nipah virus infection admitted to hospitals, diagnosed in March - May 1999, and controls, Port Dickson, Negeri Sembilan, Malaysia. No. of cases (%) No. of controls (%) Age (years)a < 20 3 5.8 4 12.9 20 - 29 7 13.5 6 19.4 30 - 39 16 30.8 3 9.7 40 - 49 16 30.8 6 19.4 > 49 10 19.2 12 38.7 Sex Male 44 84.6 19 61.3 Female 8 15.4 12 38.7 Race Chinese 39 75.0 13 41.9 Indian 11 21.2 17 54.8 Others 2 3.8 1 3.2 aMean age of cases and controls were 39.3 and 43.8 years respectively. The youngest and oldest were 14 and 77 years for cases and 15 and 78 years for controls respectively. The majority of cases were males (84.6 %) and Table 3 ages between 30 to 49 years (61.6 %), with mean Distribution of occupation for Nipah virus age of 39.3 years.