Stool specimens of 104 primary schoolchildren (mean+/-SD age = 8.2+/-0.3 years) were examined for helminth eggs and for occult blood to investigate the possibility that trichuriasis causes occult intestinal bleeding in the absence of the overt Trichuris dysentery syndrome. A commercially available guaiac test was used to detect fecal occult blood. Sixty-one children had Trichuris infection, 11 of whom had heavy infections (> 10,000 eggs per gram of feces [epg]), and 53 had Ascaris infections. No hookworm infection was detected. Baseline screening yielded only one weakly positive occult blood test result in a child with a light (800 epg) Trichuris infection. Serial stool occult blood testing on the 11 subjects with heavy trichuriasis and 8 uninfected controls yielded a single weakly positive result in the control group. The results provide no evidence that trichuriasis predisposes to significant occult gastrointestinal bleeding in children in the absence of the dysenteric syndrome.
The objective of this study was to investigate the longevity of positive dot enzyme immunosorbent assay (dot EIA) results for IgM and IgG to a Salmonella typhi outer membrane protein in Malaysian children with enteric fever. The patients were children one month to 12 years of age with clinical evidence of typhoid fever, positive blood or stool cultures for S. typhi, and/or a positive Widal test result who were admitted over a two-year period to General Hospital (Kota Bharu, Malaysia). These patients received standard inpatient treatment for enteric fever including chloramphenicol therapy for 14 days. Dot EIA tests were performed as part of clinical and laboratory assessments on admission, at two weeks, and then at 3, 6, 9, 12, 15, 18, and 21 months postdischarge. Assessment of the longevity of positive dot EIA IgM and IgG titers was done by Kaplan-Meier analysis. In 94 evaluable patients, 28% were dot EIA IgM positive but IgG negative on admission, 50% were both IgM and IgG positive, and 22% were IgM negative and IgG positive. Mean persistence of IgM dot EIA positivity was 2.6 months (95% confidence interval = 2.0-3.1 months) and that of IgG was 5.4 months (4.5-6.3 months). There were no significant differences between the three subgroups. Thus, positive IgM and IgG results determined by dot EIA within four and seven months, respectively, following documented or suspected enteric fever in a child from an endemic area should be interpreted with caution. In other clinical situations, the dot EIA remains a rapid and reliable aid to diagnosis.
A clinical trial on the efficacy of a single oral dose of ivermectin at 20, 50, 100, and 200 micrograms/kg was carried out in 40 subjects with subperiodic Brugia malayi microfilaremia. There was no significant difference in the clearance of microfilaremia in the four treatment groups, and the lowest geometric mean microfilarial count (GMC) achieved in the 40 subjects was 8.8/ml or 8.3% of the initial count (106.1/ml), at two weeks post-treatment. The GMC started to increase at one month post-treatment and by six months was 22.2% of the initial GMC. Only 27.5%, 23.1%, 15.0%, and 18.9% of subjects were amicrofilaremic at two, four, 12, and 24 weeks post-treatment, respectively. Mild fever in 35% of the subjects was the primary side reaction and was more common in those with microfilarial counts > or = 500/ml (85.7%) than in those with counts < 500/ml (32%). The clearance of B. malayi microfilaremia by ivermectin was less rapid than that reported for Wuchereria bancrofti. The smaller number of side reactions encountered in the present study compared with those reported for bancroftian filariasis is probably related to the lower microfilarial density in the present subjects. Since ivermectin at a single oral dose of 20-200 micrograms/kg can reduce the GMC to less than 10% at two weeks and maintain it below 25% of the initial level even at six months post-treatment, it is recommended that the drug be seriously evaluated for use in the control of brugian filariasis.
Two hundred forty nucleotides from the pre-membrane gene region of 12 Japanese encephalitis virus (JEV) strains isolated from three different regions of Malaysia from 1993 to 1994 were sequenced and compared with each other and with the JEV strains from different geographic areas in Asia. These 12 Malaysian isolates were classified into two genotypes. The four JEV strains isolated from Sarawak in 1994 and the four JEV strains isolated from Sepang, Selangor in 1993 were classified into one genotype that included earlier isolated strains from Malaysia (JE-827 from Sarawak in 1968 and WTP/70/22 from Kuala Lumpur in 1970). The four JEV strains from Ipoh, Perak in 1994 were classified into another genotype that included JEV strains isolated from northern Thailand and Cambodia. In an earlier report, 10 JEV strains from Sabak Bernam, Selangor in 1992 were classified into the largest genotype that included strains isolated in temperate regions such as Japan, China, and Taiwan. The data indicate that at least three genotypes of JEV have been circulating in Malaysia.
We compared a commercially available dot-blot immunoassay system with the indirect immunofluorescence assay (IFA) in tests of known negative and known positive sera from scrub typhus cases. Using a panel of 100 sera from patients with various rickettsial and nonrickettsial infections, we observed that the IFA was 99% specific and the dipstick assay was 98% specific. In tests of 91 sera (30 negative and 61 positive for scrub typhus antibodies) from a study of febrile patients in Malaysia, using the standard of an IFA titer < 1:64 as negative, an IFA titer > 1:128 as positive, and an IFA titer = 1:64 as either positive or negative (supported by clinical records), dipsticks were 83% specific and 90% sensitive. The quantitative correlation of the dipsticks to IFA titers was confirmed by significant differences in geometric means of inverse IFA titers corresponding to the number of positive dipstick spots (no dots = 8.5, one dot = 43.3, two dots = 206.7, and three dots = 676.9). The assay would enable physicians and public health workers who deal with patients to quickly diagnose and appropriately treat most cases of the disease, especially in areas of high prevalence where the proportion of false-positive results to true-positive results would be low.
In October 1988, 13 Chinese children died of acute hepatic encephalopathy in the northwestern state of Perak in peninsular Malaysia. The acuteness of the illness differed from previously reported outbreaks described in Kenya, India, and Thailand. Epidemiologic investigations determined that the children had eaten a Chinese noodle, loh see fun, hours before they died. The attack rates among those who had eaten the noodles were significantly higher than those who had not (P < 0.0001). The cases were geographically scattered in six towns in two districts along the route of distribution of the noodle supplied by one factory in Kampar town. Aflatoxins were confirmed in postmortem samples from patients. This outbreak has important public health implications for many developing countries.
We describe here a reverse transcriptase-polymerase chain reaction method for the detection of malaria parasites. Ten in vitro-cultured isolates of Plasmodium falciparum and 16 specimens from patients infected with P. falciparum were used to examine the specificity and sensitivity of the test. The sensitivity of the test was 0.3 parasites per microliter of blood. Specificity was determined by matching the sequences of the specimens' DNA to published sequences of 18S ribosomal RNA genes in the species-specific region. The test proved to be very sensitive and specific for the detection of P. falciparum infection.
A visual, enzyme-linked immunosorbent assay using urease (ELISA-U) as the enzyme marker was adapted for rapid detection of antibody against Plasmodium falciparum. Flat-bottom, 96-well microtiter plates were coated with P. falciparum soluble antigen obtained by saponin and NP-40 treatment of parasite cultures. Antibody was detected by successive incubations with test sera, urease-conjugated rabbit-human antibody, and urease substrate. Reactive sera developed a definite and easily visualized purple color. Sera from patients with single infections of P. vivax or P. ovale were unreactive. No cross-reactivity was noted with sera from patients with rheumatoid arthritis, filariasis, amebiasis, schistosomiasis, dengue, scrub typhus, leptospirosis, or toxoplasmosis. The procedure can be performed at room temperature and completed within 1 hr. The sensitivity of the assay is comparable to that of the indirect fluorescent antibody test at all but the lowest dilutions tested.
We studied 1,629 febrile patients from a rural area of Malaysia, and made a laboratory diagnosis in 1,025 (62.9%) cases. Scrub typhus was the most frequent diagnosis (19.3% of all illnesses) followed by typhoid and paratyphoid (7.4%); flavivirus infection (7.0%); leptospirosis (6.8%); and malaria (6.2%). The hospital mortality was very low (0.5% of all febrile patients). The high prevalence of scrub typhus in oil palm laborers (46.8% of all febrile illnesses in that group) was confirmed. In rural Malaysia, therapy with chloramphenicol or a tetracycline would be appropriate for undiagnosed patients in whom malaria has been excluded. Failure to respond to tetracycline within 48 hours would usually suggest a diagnosis of typhoid, and indicate the need for a change in therapy.
Serum samples were obtained within 3 days of capture from 106 cynomolgus monkeys (Macaca fascicularis) in peninsular Malaysia. Fifty-two monkeys were trapped on the fringes of palm oil estates and 54 in dense primary jungle. Sera were tested for antibodies to hepatitis A virus (HAV) with a commercial radioimmunoassay. Twenty-four animals had detectable serum anti-HAV activity (6 of 52 from palm oil estate sites and 18 of 54 from primary jungle sites). Among monkeys at both sites, antibody prevalence was strongly correlated with animal weight: overall only four of 69 monkeys (6%) weighing less than 2.0 kg had serum anti-HAV antibodies, while 14 of 29 (48%) weighing 2.0 to 3.9 kg, and 6 of 8 (75%) weighing 4.0 kg or more, had serum anti-HAV antibodies. These data suggest that wild cynomolgus monkeys in Malaysian jungles become infected with HAV or an HAV-like virus at a rate comparable to that of humans in the same region, and raise the possibility of a sylvatic cycle for HAV.
A new and rapid malaria immunoperoxidase assay using the enzyme horseradish peroxidase in place of fluorescein isothiocyanate was developed to allow the serological measurement of antimalarial antibody by light microscopy. Acetone-fixed thin blood films prepared from cultured Plasmodium falciparum were used as the source of antigen. This malaria immunoperoxidase assay is as sensitive as, and occasionally more sensitive than, the indirect fluorescent antibody assay. It is easy to perform and the antigen used does not show cross-reactivity with sera from nonmalarial diseases.
Dexamethasone has recently been shown to block the production of cachectin (implicated in the pathogenesis of cerebral malaria) if administered prior to endotoxin induction of mouse macrophages. Using the hamster cheek pouch-cerebral malaria model, we tested the hypothesis that dexamethasone is effective as a therapeutic agent in severe malaria if given before some yet undefined trigger point in the disease. Infected hamsters were treated with dexamethasone (0.7 mg/kg) daily on days 7-12, 4-12, or 1-12 post-challenge. When treatment was started on day 1, whole body oxygen consumption (used as a measure of erythrocyte transport to sites of diffusion) on day 12 was greater than (P less than 0.05) that of infected control animals, though the degree of anemia was no different in treated and untreated groups. Furthermore, treatment produced a reduction in monocyte accumulation, capillary malfunction, and monocyte/red blood cell aggregate formation observable in the cheek pouch in vivo and a similar reduction in monocyte presence, capillary pathologic change, and multifocal hemorrhage in the brain on postmortem. These data suggest that mediator(s), whose production can be blocked by pretreatment with dexamethasone, are involved in the pathogenesis of disease leading to death of the Plasmodium berghei infected hamster.
The sensitivities and specificities of the indirect microimmunofluorescent antibody (IFA) and Weil-Felix (OXK) tests for scrub typhus were established for a range of titers using groups of diseased and control (other febrile illnesses) patients diagnosed by other methods. At a cut-off point of greater than or equal to 1:400, the IFA test was 0.96 specific, and at greater than or equal to 1:320, the OXK was 0.97 specific. Using either these highly specific levels of antibody or other rigorous diagnostic criteria (isolation or 4-fold rising titers), the prevalence of scrub typhus infection was determined to be 0.22 in an unselected population of febrile patients in a rural Malaysian hospital. Probability values (Pr) for the correct diagnosis of scrub typhus were then calculated from the specificity, sensitivity and prevalence determination for a range of titers. The Pr for an OXK titer of greater than or equal to 1:320 was 0.79, and the Pr for an IFA titer of greater than or equal to 1:400 was 0.78. When both these titers were present in a single specimen, the Pr increased to 0.96.
Albendazole, a new anthelmintic drug was evaluated in Malaysia in 91 patients, with single or mixed infections of Ascaris, Trichuris, and hookworm. Albendazole was administered as a single dose of 400 mg, 600 mg, or 800 mg. The cure rate for Ascaris at all three doses was 100% at days 14 and 21 post-treatment; for hookworm it was 98.8%, 100% and 98%, respectively, at day 14 and 68.8%, 100% and 84%, respectively, at day 21; for Trichuris it was 31.2%, 57.1% and 42.3%, respectively, at day 14 and 27.3%, 60.9% and 48.0%, respectively, at day 21. The egg reduction rate at day 21 was 100% at all three doses for Ascaris, 94.5%, 100% and 96.1%, respectively, for hookworm; and 39.2%, 85.1% and 72.8%, respectively, for Trichuris. There were no side effects, and biochemical examination of blood and urine did not indicate any unfavourable changes. Based on this trial, the recommended dosage for Ascaris and hookworm is a 400 mg single dose, and for Trichuris is a 600 mg single dose. Albendazole appears to be more effective than other available anthelmintic drugs.
Monoclonal antibodies (MAbs) directed against different epitope regions on three sexual stage-specific gamete surface proteins of Plasmodium falciparum, Pfs 25, Pfs 230, and Pfs 48/45, were used to study the genetic diversity of these epitopes among fresh isolates of P. falciparum from Malaysia, using immunofluorescence microscopy (IFA). Among 45 Malaysian isolates, one epitope of Pfs 25, designated region I, showed evidence of variable reactivity with MAbs among different isolates; the Pfs 25 epitope, region II, was universally recognized by MAbs in all isolates. Two apparently distinct epitope regions of Pfs 230 were defined by MAbs, one of which was universally recognized by MAbs among the 45 isolates; the other was conserved in all but three isolates. The epitope regions of gamete-surface protein Pfs 48/45, designated regions I, IIa, IIb, IIc, III, and IV, were examined for reactivity by IFA in 33 isolates. Epitope regions I, IIb, III, and IV were conserved in all isolates; regions IIa and IIc existed in variant forms.
A seroepidemiologic survey of Plasmodium vivax and Plasmodium falciparum transmission was conducted in 94 Orang Asli children and adults. The prevalence of malaria was 46% in this population, and infections due to P. vivax and P. falciparum occurred with equal frequency. Multi-species infection was common, particularly in children less than 10 years of age. Circumsporozoite (CS) antibodies to P. vivax were detected by ELISA, using the recombinant protein NS181V20, in sera from 53-95% of all subjects in this study. The specificity of reactivity to NS181V20 was confirmed by immunofluorescence using air-dried sporozoites. CS antibodies to P. falciparum were present in less than 50% of the population less than 30 years of age. These data support further testing of this protein as a candidate vivax vaccine.
Mass drug administration via 3 modes of delivery reduced the incidence and prevalence rates and intensity of Brugia malayi infection in 3 rural villages in the Bengkoka Peninsula, Sabah, in 1982-1983. A dosage of 6 mg diethylcarbamazine citrate (DEC-C)/kg body weight was administered either daily or weekly (total of 6 doses, 36 mg/kg body weight), and impact on B. malayi cases were comparable in the 3 villages. A total of 384 people participated in the DEC-C regimens, and all pregnant women and children under 2 years were excluded from the study. Bekessy's method of estimation of incidence and recovery rates was applied to data on B. malayi microfilaremia before drug administration. Treatment with DEC-C by any of the 3 modes of delivery drastically reduced the number of episodes of patent microfilaremia, incidence and prevalence, and median microfilarial density. Reduction was sustained for at least 18 to 24 months after treatment.
Electrophoretically-detected allozyme variation is described in strains of Schistosoma japonicum (4 Philippine strains), S. mekongi (Laos), and an undescribed anthropophilic S. japonicum-like schistosome from Peninsular Malaysia. Result, together with those reported previously for 8 other strains (S. japonicum, China, Formosa, Japan, Philippines; S. mekongi, 2 substrains; Malaysian schistosome, 2 strains) permit a composite genetic characterization of 15 strains of Asian schistosomes at 9-18 presumptive loci. The proportion of polymorphic loci (P) and the mean heterozygosity per locus (H) were zero in all strains. Although this was expected for strains that had been in laboratory culture for up to 50 years, we expected to detect variation in strains based on 10-50 recently field-collected infected snails. We expected S. japonicum to be as variable as S. mansoni (P = 0.13 (0-0.33), H = 0.04, 18 loci, 22 strains) as it is believed to reproduce sexually, has an evolutionary history of several million years, inhabits a wide geographic range, coevolved with a genetically variable intermediate snail host, and has a diversity of mammalian hosts. No differences were detected between the 5 S. japonicum strains from Leyte and Luzon (Philippines), between the 3 S. mekongi strains, or between the 3 Malaysian schistosome strains; these groups and the remaining S. japonicum strains representing Mindoro (Philippines), China, Formosa, and Japan each have distinctive multilocus electromorphic patterns. Nei's genetic distances (D) were calculated to estimate interstrain and interspecific divergence. Interstrain genetic distances in S. japonicum averaged greater than 0.3; much higher than those reported previously for S. mansoni (D = 0.06, D(max) = 0.24). S. japonicum (Mindoro) was moderately differentiated from the Leyte-Luzon strains (D = 0.29, 12 loci). Estimates of the S. japonicum China-Philippine distance (D greater than 0.4, 11 loci) are high for conspecific populations and further studies of the still poorly characterized Chinese parasite may reveal that these are, in fact, separate species. S. japonicum is shown to be only distantly related to S. mekongi and the Malaysian schistosome (D greater than 1); the latter is closely related to, but genetically quite distinct from, S. mekongi (D = 0.61 +/- 0.275, 11 loci) and warrants recognition as a new species. The medical significance of the isogenic nature of the Asian schistosome strains and their evolutionary divergence are discussed.
A survey of 308,101 primary school children in Peninsular Malaysia conducted in 1979 by the School Health Services, Ministry of Health, Malaysia, revealed that 10.7% of children were infested with Pediculus humanus capitis. The prevalence rate was higher in the economically less advanced states of Terenganu (34%), Kelantan (23%), and Perlis (21%) than in the other states (4-13%). Of 14,233 school children examined in the State of Melaka, 26% of Indians, 18.7% of Malays, 6.1% of Europeans, and 0.7% of Chinese had pediculosis. The prevalence rate, which has remained unchanged over the past 5 years, does not appear to vary with age but is higher in children with long hair and those from the lower socioeconomic groups. Boys have a lower infestation rate than do girls. The higher incidence in Indians and Malays correlates well with their lower socioeconomic status in the community, and their cultural habit of maintaining longer hair than do the Chinese. The difference become less apparent in the higher socioeconomic groups.
Scrub typhus is a major cause of febrile illness throughout the Asia-Pacific region. It is commonly undiagnosed, partly because of the lack of a simple, reliable diagnostic test which can be used in clinical laboratories. The indirect immunoperoxidase technique, configured into a test kit, was provided to technicians who were trained in its use. They used the kit during a 2 year field trial in their respective clinical hospital laboratories throughout Malaysia. In an evaluation using 1,722 consecutive sera tested in those laboratories, the kit was found to have a median sensitivity for IgG detection of 0.85 (range 0.33-0.95), a median specificity of 0.94 (range 0.88-1.00), reproducibility of 0.86, and efficiency of 0.92 when compared to the reference laboratory. In a proficiency survey in which 10 laboratories received 3 coded test samples, all but 2 laboratories had results within 1 dilution of the reference laboratory in quantitating specific IgG, whereas 7 laboratories were within 1 dilution in quantitating IgM. The shelf life of the kit was at least 1 year at 4 degrees C.