Inhibition of lymphocyte proliferative responses to Helicobacter pylori by plastic adherent cells.

A study was carried out on 49 H. pylori-positive and 11 H. pylori-negative patients to determine the reactivity of peripheral blood lymphocytes (PBL) to phytohemagglutinin (PHA) and acid glycine extract (AGE) of H. pylori, and to identify cells responsible for imunosuppression. Based on response to PHA stimulation, cell-mediated immunity of all patients were competent. In some patients, however, response to AGE of H. pylori was suppressed by plastic adherent cells. This study provided evidence of the presence of plastic adherent suppressor cells which suppressed PBL response to AGE of H. pylori but not to PHA suggesting that immunosuppression is antigen specific. There is also an indication that immunosuppression may be species-specific as PBL devoid of plastic adherent cells only responded to stimulation by AGE of H. pylori but not that to AGE of C. jejuni.

Toxocara seroprevalence and childhood asthma among Malaysian children.

BACKGROUND: The larva of Toxocara spp., a common animal roundworm, may infect non-compatible hosts, causing a profound immunological reaction with marked eosinophil and IgE responses, not unlike in atopy. In this study, we determined the seroprevalence of Toxocara exposure in 66 asthmatic and 58 non-asthmatic children. METHODS: Exposure to Toxocara was determined by examining the serum samples of the children for specific IgG antibodies to L2 Toxocara larvae, using a commercially available diagnostic kit. RESULTS: There was no significant difference in the mean age, sex, social class, residence type and presence of domestic pets at home between the two children groups. Children with bronchial asthma were observed to have higher Toxocara seropositivity than that of the non-asthmatic controls (21.2 vs 8.6%, P=0.047). CONCLUSION: The observed relationship between exposure to Toxocara infection and bronchial asthma in Malaysian children warrants further evaluation. An understanding of any possible contribution to the pathogenesis of childhood asthma provides a potential avenue for prevention.

Specificity and sensitivity of a rapid dipstick test (Brugia Rapid) in the detection of Brugia malayi infection.

A total of 753 serum samples from 6 institutions in 3 countries (Malaysia, Indonesia and India) were used to evaluate an immunochromatographic rapid dipstick test, Brugia Rapid, for diagnosis of Brugia malayi infection. The samples comprised sera from 207 microfilaria-positive individuals and 546 individuals from filaria non-endemic areas. The latter consisted of 70 individuals with soil-transmitted helminth infections, 68 with other helminth infections, 238 with protozoan infections, 12 with bacterial and viral infections and 158 healthy individuals. The dipstick is prepared with a goat anti-mouse antibody control line and a B. malayi recombinant-antigen test line. First, the dipstick is dipped into a well containing diluted patient serum, thus allowing specific anti-filarial antibody in the serum to react with the recombinant antigen. Then the dipstick is placed into an adjacent well containing reconstituted anti-human IgG4-gold. After 10 min, development of 2 red-purplish lines denotes a positive result and one line indicates a negative reaction. The overall results of the evaluation showed 97% sensitivity, 99% specificity, 97% positive predictive value and 99% negative predictive value. Brugia Rapid is thus a promising diagnostic tool for detection of B. malayi infection, and would be especially useful for the brugian filariasis elimination programme.

The prevalence of malnutrition and geo-helminth infections among primary schoolchildren in rural Kelantan.

A cross-sectional study of the nutritional status of schoolchildren aged 7-10 years from the Kuala Betis Resettlement Scheme in Gua Musang, Kelantan was done. A total of 291 schoolchildren were examined but only 183 (62.9%) fecal samples were returned for geo-helminth infection analysis. The prevalence of stunting was 40.4% and underweight was 28.4%. A total of 127 schoolchildren (69.4%) were positive for at least one of the 3 geo-helminth infections. The most common geo-helminth was Ascaris lumbricoides (62.8%), followed by Trichuris trichiura (38.9%), and hookworm infection was relatively low (12.6%). All the anthropometric indices measured were lower in the geo-helminth infected children compared to the uninfected childen, except for the weight-for-height z-score. However the differences were insignificant Geo-helminth infections may not be a significant factor in malnutrition of these schoolchildren. However, with the availability of safe, efficacious and cheap broad spectrum anthelmintics, regular mass treatment should be given in selected areas where the prevalence of geo-helminth infections is still high, so that these schoolchildren will be able to achieve their growth potential during their school years.

Field's stain--a rapid staining method for Acanthamoeba spp.

Acanthamoeba sp. is a free-living amoeba known to cause chronic central nervous system infection or eye infection in humans. Many cases remain undetected for want of a good detection system. We report for the first time a rapid staining method to facilitate the identification of Acanthamoeba sp. using the modified Field's staining technique. A. castellanii, which was used in the present experiment, is maintained in our laboratory in mycological peptone medium (Gibco). The cultures were pooled together and smears were made on glass slides for staining purposes. Different types of stains such as Field's stain, modified Field's stain, Wright's stain, Giemsa stain, Ziehl-Neelsen stain, and trichrome stain were used to determine the best stain for the identification of this amoeba. The concentration of various stains and the duration of staining were varied to provide the best color and contrast for each stain. Acanthamoeba was also obtained from the brain of experimentally infected mice and was stained with various stains as mentioned above to determine the best stain for use in identifying the presence of this parasite in experimentally infected animals. The modified Field's stain gives a very good color contrast as compared with other stains. Furthermore, it takes only 20 s to be carried out using the least number of reagents, making it suitable for both laboratory and field use.

PCR-ELISA for the detection of Brugia malayi infection using finger-prick blood.

A polymerase chain reaction assay based on the enzyme-linked immunosorbent assay (PCR-ELISA) has been developed to detect Brugia malayi infection in an area of low endemicity in Malaysia. Blood samples from 239 subjects were tested: 192 amicrofilaraemic individuals, 14 microfilaraemic persons and 3 chronic elephantiasis cases from endemic areas and 30 city-dwellers (non-endemic controls). PCR products were examined by ELISA and Southern hybridization. In the PCR-ELISA, digoxigenin-labelled PCR products were hybridized to a biotin-labelled probe. This was followed by incubation in streptavidin-coated microtitre wells and detection using anti-digoxigenin-peroxidase and ABTS [2,2'-azinobis(3-ethylbenzthiazoline-6-sulphonic acid)]. All microfilaraemic samples were positive by PCR-ELISA and Southern hybridization and all samples from non-endemic subjects and chronic elephantiasis patients were negative. The PCR-ELISA detected 12 times as many B. malayi infections as did thick blood film examination. Nineteen of the 194 samples from the endemic area gave positive results by both PCR-ELISA and Southern hybridization, and an additional 5 samples were positive by PCR-ELISA only. The PCR-ELISA was specific and sensitive, detected more infections, and was more reproducible than Southern hybridization.

A Brugia malayi antigen specifically recognized by infected individuals.

Western blot analyses were performed on 444 serum specimens: 40 sera from microfilaraemic individuals, 10 sera from elephantiasis patients, 24 treated individuals, 50 sera from residents of endemic areas without anti-filarial IgG4 antibodies (endemic normals), 20 sera from amicrofilaraemic individuals with high anti-filarial IgG4 antibodies, 200 sera from healthy city-dwellers (non-endemic samples), and 100 sera from soil-transmitted helminth-infected individuals. Phast electrophoresis system was used to electrophorese Brugia malayi soluble adult worm antigen on 10-15% SDS-PAGE gradient gels followed by electrophoretic transfer onto PVDF membranes. Membrane strips were then successively incubated with blocking solution, human sera, and monoclonal anti-human IgG4 antibody-HRP, with adequate washings done in between each incubation step. Luminol chemiluminescence detection was then used to develop the blots. An antigenic band with the MW of approximately 37 kDa was found to be consistently present in the Western blots of all microfilaraemic sera, all amicrofilaraemic sera with high titres of anti-filarial IgG4 antibodies, some treated patients, and some elephantiasis patients. The antigen did not occur in immunoblots of individuals with other helminthic infections, normal endemic individuals, and city dwellers. Therefore the B. malayi antigen of with the MW of approximately 37 kDa demonstrated specific reactions with sera of B. malayi-infected individuals and thus may be useful for diagnostic application.

Use of antifilarial IgG4-ELISA to detect Brugia malayi infection in an endemic area of Malaysia.

OBJECTIVE: To evaluate the usefulness of antifilarial IgG4 antibody assay in detecting B. malayi infection in a filaria endemic area in Malaysia. METHODS: A sandwich ELISA using B. malayi soluble antigen was employed to detect antifilarial IgG4 antibodies in serum samples of 330 individuals who comprised 88 healthy individuals from nonendemic areas, 15 B. malayi microfilaraemic cases, 22 individuals with soil-transmitted helminthiases, 9 elephantiasis cases and 196 residents from a B. malayi-endemic area. An O.D. value of > 0.420 at serum dilution of 1:400 was used as the cut-off point. This cut-off point was obtained by taking the mean optical density (0.252 + 4 S.E.) of 36 negative sera which had O.D. values greater than 0.1 at serum dilution of 1:400. RESULTS: All 15 microfilaraemic persons were positive for antifilarial IgG4 antibody. Non-endemic normals, soil-transmitted helminth infected persons and chronic elephantiasis cases were negative for antifilarial IgG4 antibody. Of the 196 individuals from the filaria endemic area, 37 (18.8%) demonstrated presence of antifilarial IgG4 antibodies; and only eight individuals (4.1%) were positive for microfilariae. All eight microfilaraemic individuals were also positive for antifilarial IgG4 antibodies. CONCLUSION: Antifilarial IgG4-ELISA could detect 4.6 times more positive cases than the microfilaria detection method. With appropriate cut-off values that eliminate cross-reactivities, this serological tool is very useful for Brugia malayi prevalence surveys and diagnosis.

Imported malaria: a retrospective study in University Hospital, Kuala Lumpur, a ten-year experience.

Over a period of ten years (1983-1992), 134 malaria cases admitted to University Hospital, Kuala Lumpur (UHKL) were analysed. Malays constituted 27.6%, Chinese 29.8%, Indians 9.7%, Indonesians 16.4% and other foreigners 16.4%. Therefore, of the total number of cases, foreigners constituted 32.8% (44) of all the malaria cases admitted to UHKL. Fifteen of these foreigners had chloroquine-resistant strains of malarial parasites. Three species of malaria were reported of which Plasmodium falciparum constituted the most (46.3%) (80% of these developed resistance to chloroquine). Plasmodium vivax was confirmed in 44.8% (10% of these developed resistance to chloroquine) and there was only one case of Plasmodium malarie infection.

Cor pulmonale: an unusual presentation of tropical eosinophilia.

Tropical pulmonary eosinophilia (TPE) is considered to be a variant of human filarial infection. The pulmonary manifestations of TPE have been well described. Extra-pulmonary features of the disease, although not commonly seen, have been reported previously. A 9-year-old Malay girl with a history of recurrent cough and wheezing was admitted because of cardiac failure. Physical examination revealed a very sick girl with tachypnoea, central cyanosis, finger clubbing, elevated jugular venous pulse, generalized crackles and rhonchi in the chest, a loud second heart sound and hepatosplenomegaly. A chest radiograph showed cardiomegaly and right pleural effusion. Laboratory investigations revealed hypochromic, microcytic anaemia with persistent blood eosinophilia (absolute eosinophil counts varied from 1.9 to 5.5 x 10(9)/1). The ELISA test for antifilarial IgG antibodies was strongly positive. She responded promptly to treatment with diethylcarbamazine. In summary, this is a patient with TPE who presented with cor pulmonale, probably due to late-stage interstitial pulmonary fibrosis. In order to prevent the long term morbidity of cardiorespiratory disability, the early signs of TPE should be recognized and the infection treated.

Effect of intestinal helminthiasis on school attendance by early primary schoolchildren.

Stool examination of 249 early primary schoolchildren at 2 schools in north-eastern peninsular Malaysia revealed that 73 were infected with Ascaris lumbricoides, 103 with Trichuris trichiura, and 18 with hookworms. Infected children were treated with a single dose of 400 mg of albendazole. The school attendance records during a 60 d period before treatment and 2 consecutive 60 d periods after treatment were examined. The absenteeism rate did not improve more among infected children after treatment than it did among the uninfected control children. The correlation between worm intensity and the number of lost school-days was poor. There was no evidence that intestinal helminthiasis caused school absenteeism among this group of children.

Effect of intestinal helminthiasis on intestinal permeability of early primary schoolchildren.

Intestinal permeability of 246 early primary schoolchildren at 2 schools (106 of whom were infected with intestinal helminths) was assessed by using the lactulose/mannitol differential absorption test. The ratio of the urinary recoveries of lactulose and mannitol was determined after oral administration of a standard solution of the 2 sugars. Assessment of intestinal permeability was repeated on 100 infected children after treatment and on a cohort of 68 uninfected children. Infected and uninfected groups were compared with respect to baseline lactulose/mannitol ratio (L/M1) and change in lactulose/mannitol ratio between assessments (delta L/M). The correlations between baseline intensity of infection and L/M1, and between fall in intensity and delta L/M, were evaluated. Based on a crude index of socioeconomic status, each child was assigned to one of 3 socioeconomic groups; all but 3 children belonged to either groups 2 or 3. Trichuris trichiura and Ascaris lumbricoides were the 2 predominant infections; the hookworm infection rate was relatively low. The results suggested that helminthiasis exerted only a marginal effect on intestinal permeability, the impact of which in children from lower socioeconomic backgrounds was negligible in comparison with the cumulative effects of other factors.

Potential use of IgG2-ELISA in the diagnosis of chronic elephantiasis and IgG4-ELISA in the follow-up of microfilaraemic patients infected with Brugia malayi.

Sera from fifty subjects with different presentations of Brugian filariasis and from common soil-transmitted helminth infections were tested for specific anti-filarial IgG and its subclasses. Anti-filarial IgG, IgG1 and IgG3 showed cross-reactivities with soil-transmitted helminthic infections and no significant differences in optical densities among the various groups of filarial patients. In comparison with other groups of subjects, IgG4-ELISA of sera from microfilaraemic patients and some previously microfilaraemic patients showed a significant increase in optical density readings, while IgG2-ELISA showed elevated optical density readings in sera of patients with chronic elephantiasis. Therefore IgG2-ELISA is potentially useful in the diagnosis of brugian chronic elephantiasis while IgG4-ELISA may be beneficial for follow-up diagnosis of treated microfilaraemic patients.

Reliability of two commercial serological kits for serodiagnosing Helicobacter pylori infection.

Two commercial serological kits, Pylori-set (Orion Diagnostica, Finland) and Helico-G (Cambridge Biomedical Ltd, UK), and an in-house ELISA were evaluated with sera from 24 Helicobacter pylori-positive and 146 H. pylori-negative dyspeptic patients. Sensitivity, specificity, positive and negative predictive values of Pylori-set were lower than that of Helico-G and in-house ELISA. Helico-G was more sensitive (91.7%) than in-house ELISA (83.3%) and both had comparable negative predictive values of 98.3% and 97.3%, respectively. However, specificity (97.9%) and positive predictive value (86.9%) of an in-house ELISA were much higher than specificity (80.1%) and positive predictive value (43.1%) of Helico-G. Kappa index of agreement between the three serological tests (Pylori-set, Helico-G or in-house ELISA) and the presence of H. pylori in antral biopsies was very low (k = 0.13; z = 1.9; p > 0.05), moderate (k = 0.49; z = 7.1; p < 0.0001), or substantial (k = 0.82; z = 10.8; p < 0.0001), respectively. Overall, statistical evaluations demonstrated that both commercial kits were not as reliable as the in-house ELISA for serodiagnosing H. pylori infection.

Helicobacter pylori infection in north-eastern peninsular Malaysia. Evidence for an unusually low prevalence.

The prevalence of Helicobacter pylori infection was determined in peptic ulcer patients, in non-ulcer dyspepsia (NUD) patients, and in the general adult population. The H. pylori infection rate ascertained by microbiologic examination of multiple gastric antral biopsy specimens was 50% (17 of 34) in duodenal ulcer (DU), 5% (1 of 22) in gastric ulcer, and 9% (15 of 159) in NUD patients. A seroepidemiologic survey showed a prevalence of only 4.2% among 496 blood donors and 4.8% among 921 subjects who attended health screening clinics. H. pylori infection is relatively uncommon and does not appear to be the predominant factor in the pathogenesis of peptic ulcer disease in the area. The incidence of peptic ulcer perforations in the area in 1991-92 was 1.5 per 100,000 person-years, reflecting a relatively low frequency of peptic ulcers, which might be due to the low prevalence of H. pylori infection in the population.

Demonstration of antigenic similarities and variations in excretory/secretory antigens of Toxoplasma gondii.

C57BL/6 mice were orally infected with different doses of cysts of ME49 strain of Toxoplasma gondii to produce groups of acutely and chronically infected mice. Sera were obtained at different periods post-infection. SDS-PAGE was ran with excretory/secretory antigens of ME49 and RH strains of T. gondii, followed by Western blot analyses using the above sera and anti- IgA, IgM, IgG as conjugates. The SDS-PAGE profiles of the two antigens were similar. However the antigenic bands showed variations in all blots, most evidently in IgA blots of chronic sera. IgG blots showed greatest similarities in reactive bands. In IgM blots, more common bands were shown in chronic sera than in acute sera. Variations and similarities in prominence of some bands and time of their appearance were also noted, especially in IgM and IgG blots of chronic sera. Thus antigenic variations and similarities are present in excretory/secretory products of different strains of T. gondii.

Cryptosporidiosis among children with acute gastroenteritis in the pediatric ward in the General Hospital, Penang.

Stool samples from 836 cases with diarrhea and acute gastroenteritis from the Pediatric ward, Penang General Hospital, were examined for Cryptosporidium oocysts. A dimethyl sulfoxide modified acid fast technique was used for the identification of the parasites. 36 samples or 4.3% were found to be positive for Cryptosporidium. The prevalence of infection was higher (2.39%) in children with diarrhea and vomiting than in children with acute gastroenteritis alone (0.8%). Stool examination and cultures from the Cryptosporidium positive samples revealed no other parasites, rotavirus or enteropathogenic bacteria. This suggests that Cryptosporidium may be an important agent in the causation of diarrhea in young children. A routine laboratory examination for the detection of Cryptosporidium in the search for causal agents of childhood diarrhea in our environment may, therefore, be significant.