A survey of Nipah virus infection among various risk groups in Singapore.

Following the Nipah virus (NV) outbreak in March 1999 in Singapore, a serological survey was undertaken to screen individuals potentially exposed to NV. Blood samples were tested for NV IgM, IgG and neutralizing antibodies. Twenty-two (1.5%) of 1469 people tested had antibodies suggesting NV infection. Although 12 of the 22 infected people (54.6%) were symptomatic, the remaining 10 (45.4%) were clinically well and had no past history of compatible pulmonary or neurological disease. Clinical and serological findings suggested three people had been infected with NV before the outbreak was recognized. All those who were infected were male abattoir workers. None of the people who had contact with horses, and no healthcare workers exposed to infected patients and their specimens had detectable antibodies. This study provides evidence that NV causes asymptomatic infection. All of the antibody positive individuals had direct contact with pigs and there was no evidence of human to human transmission.

Evaluation of Amplicor- and IS6110-PCR for direct detection of Mycobacterium tuberculosis complex in Singapore.

One hundred and seventy-eight samples from 168 individuals were tested for Mycobacterum tuberculosis complex (Mtc) using Amplicor PCR, IS6110-PCR (in-house), acid fast (AF)-staining and culture. Thirty-one samples were positive by culture, but 37 samples were later resolved to be truly positive for Mtc. Of these, Amplicor detected 32 (86.5%), IS6110-PCR detected 31 (83.6%), and AF-staining 21 (56.8%). None of the 141 Mtc-negative samples was positive by these tests, thus giving 100% specificity. Although the IS6110-PCR was more sensitive than Amplicor in detecting spiked Mtc DNA, it was not more sensitive than the latter in detecting Mtc in clinical samples. Reasons likely to account for the PCR false negativity were (i) sample inoculum size, (ii) nonuniform samples due to clumping effect of Mtc and (iii) the absence of target gene sequences for IS6110-PCR. Culture negativity, on the other hand, was likely to be associated with nonviable Mtc. Amplicor PCR is promising for direct detection of Mtc. The IS6110-PCR, however, may not be as suitable because of possible existence of IS6110-deleted Mtc strain in Singapore.

PCR as a confirmatory test for human immunodeficiency virus type 1 infection in individuals with indeterminate western blot (immunoblot) profiles.

A combination of two PCR systems which amplify human immunodeficiency virus type 1 (HIV-1) gag and pol DNA was used to elucidate the HIV-1 infection status of a group of individuals screening positive for anti-HIV but having indeterminate Western blot (immunoblot) profiles. The results obtained suggest PCR could be used to resolve such cases.

Anti-toxoplasma serotitres in uveitis and ocular toxoplasmosis.

This study was based on anti-toxoplasma serotitres obtained from 80 normal healthy adults and 103 consecutive patients with anterior, posterior and panuveitis over a 42 month period. Twelve out of 80 (15%) normal healthy sera were found to have low titres (1:64 to 1:256), while 3 (3.8%) had high titres (1:1024 or higher). The distribution of serotitres in patients with clinical toxoplasmosis was significantly different from the population sample (p less than 0.001, chi-squared test). Similarly, the distribution of serotitres in patients with posterior uveitis (of which 28 out of 54 were clinically ocular toxoplasmosis) having serotitres of 1:1024 or greater, was found to be significantly different from the normal population (p less than 0.001, Chi-squared test), but not statistically different from those with clinical ocular toxoplasmosis. Hence, anti-toxoplasma serotitre is useful as a diagnostic screening tool only in patients with posterior uveitis. It is also useful in confirming cases of suspected clinically positive ocular toxoplasmosis in our local population.

Distribution of the 3.05-Mdal "Toronto" beta-lactamase plasmid among penicillinase-producing isolates of Neisseria gonorrhoeae in the Far East.

One hundred and fifty-two beta-lactamase-producing Neisseria gonorrhoeae isolates from Japan (n = 25; 1983, 1985, 1986), Taiwan (n = 14; 1983, 1984), and the Republics of Singapore (n = 58; 1986, 1987) and the Philippines (n = 55; 1989) isolated from 1983 through 1989 were characterized by auxotype, serovar, and plasmid content to determine the distribution and diversity of penicillinase-producing N. gonorrhoeae (PPNG) strains possessing the 3.05-Mdal "Toronto" beta-lactamase plasmid. PPNG isolates possessing a 3.05-Mdal beta-lactamase plasmid were isolated in Japan (1/25: 4%), Taiwan (4/14: 29%), and the Republic of the Philippines (3/55: 5%); no PPNG isolates with the 3.05-Mdal plasmid were isolated in the Republic of Singapore. All isolates possessing the 3.05-Mdal plasmid also possessed a 24.5-Mdal conjugative plasmid and belonged to the auxotype/serovar class, Proto/IB-1. Studies with five isolates possessing the 3.05-Mdal plasmid, and representing isolates from each country in which they were found, confirmed that the beta-lactamase plasmid in these strains could not be transferred to another gonococcal isolate by conjugation. PPNG isolates possessing the "Toronto" plasmid are widespread in the Far East; spread of these isolates may, however, be limited to the physical spread of a single strain.

Characterisation of PPNG and non-PPNG Neisseria gonorrhoeae isolates from Singapore.

OBJECTIVE: To characterise Neisseria gonorrhoeae isolates from Singapore. DESIGN: Characterisation of Neisseria gonorrhoeae isolates by auxotyping, serological analysis and plasmid profile analysis. SPECIMENS: Sixty randomly collected isolates from 41 symptomatic, untreated males and 19 female prostitutes were studied. RESULTS: Auxotyping of 25 PPNG and 35 non-PPNG strains showed that the Pro-auxotype was prevalent among both PPNG (56%) and non-PPNG (42.5%) strains. Prototrophic strains comprised 28% of PPNG and 32.5% of non-PPNG strains respectively. Serovar analysis showed that with the exception of seven serogroup WI strains, the majority belonged to serogroup WII/III. Serovar Aedih was predominant among both serogroup WI PPNG (80%) and non-PPNG (100%) strains. Serogroup WII/III PPNG strains were represented by nine serovars with the predominant serovars being Bacjk (28%) and Bcgjk (16%). Eleven serovars were identified in the WII/III non-PPNG strains and the major serovars were Bajk (20%), Bacjk (17%), Back (11.4%) and Beghjk (11.4%). Analysis of the 25 PPNG strains showed that 16 of them carried the 4.4 MDa (Asian type) resistance plasmid and nine strains harboured the 4.4 MDa plasmid in conjunction with the 24.5 MDa transfer plasmid. The cryptic plasmid of 2.6 MDa was present in 27 of the 35 non-PPNG strains. Five of the non-PPNG strains harbouring the cryptic plasmid also contained the 24.5 MDa transfer plasmid. The plasmid combination of 2.6 + 7.8 + 24.5 MDa was detected in three non-PPNG strains. CONCLUSION: The combination of epidemiological methods used in this study indicated the heterogeneity of N gonorrhoeae strains in Singapore. A total of 16 different combinations of auxotype, plasmid profile and serovar were seen in the 25 PPNG strains compared with 24 such combinations in the 35 non-PPNG strains. Such sensitive differentiation would otherwise not be possible using either auxotype-serovar (A/S) or auxotype-plasmid analysis.

Four patients in Singapore with anti-Golgi antibodies.

Serum specimens for anti-nuclear fluorescence tests are routinely received in our laboratory. Four specimens were spotted to be negative for anti-nuclear fluorescence but positive for fluorescence characteristic of that caused by anti-golgi antibodies. (a) Patient A had acute glomerulonephritis; (b) Patient B had acute viral hepatitis; (c) Patient C had deep vein thrombosis; and (d) Patient D had non-Hodgkin's lymphoma. The relevance and possible aetiology of anti-golgi antibodies are also discussed.

Anti-toxoplasma antibodies in healthy adults and in different patient categories.

This study analyzes the anti-toxoplasma sero-titres and prevalence rates in normal healthy adults and in patients presenting with different symptom-complexes. The study was based on sera from 80 normal healthy adults and 2,185 patient sera samples from 2,032 patients (from various clinics and hospitals in Singapore) being investigated for the diagnosis or exclusion of toxoplasmosis, over a 42-month study period. About 15% of the healthy adults were found to have low IgG antibody titres (1:64 to 1:256), while 3.8% had high IgG titres (1:1024 or higher). Interestingly, among the patients investigated for toxoplasmosis: i) more than 20% of those presenting with lymphadenopathy (usually cervical) had antibody titres more than or equal to 1:1024; ii) more than half of all patients with an antibody titre of 1:4096, and more than three-quarters of all patients with a titre exceeding 1:4096 had presented with lymphadenopathy; iii) about 20% of those presenting with ocular symptoms had low antibody titres of 1:64 or 1:256, whereas 7% had higher titres. Malay (p less than 0.01) and Indian (p less than 0.05) patients had significantly higher seropositive rates than the Chinese. In particular, the Malays (p less than 0.00001) and Indians (p less than 0.01) had significantly higher incidence of low-positive titres (1:64, 1:256); conversely the Chinese patients had a significantly higher (p less than 0.01) incidence of high-positive titres (1:4096 or higher). Finally, the epidemiology and clinical profiles of patients presenting with acute toxoplasmic lymphadenitis are contrasted with that of patients presenting with ocular manifestations of congenital toxoplasmosis.

Evidence of HTLV-I infection in Singapore prostitutes.

Sera obtained in 1987 from 63 male and 632 female Singapore prostitutes were screened for antibody to human T-cell leukaemia virus (HTLV)-I with a particle agglutination test. Of the 3 males and 4 females who were positive one had antibody to HTLV-I core and envelope antigen on Western Blot. Two subjects had presumptive antibody to HTLV-I core antigen and a third subject had such antibody on a repeat specimen in 1989. These sera were negative for HIV-1 antibody. There is evidence of infection with HTLV-I or a variant virus in this population. The infection is likely to have been sexually transmitted.

Human immunodeficiency virus infection in Singapore--containment strategies.

The worldwide occurrence of the Acquired Immunodeficiency Syndrome (AIDS) since its advent in the United States in 1981 has resulted in a global dilemma on how best to reduce transmission of the Human Immunodeficiency Virus (HIV). In Singapore, a national AIDS programme was established in 1985 to control the spread of the disease in the country. The programme included health care delivery, education of health care workers and the community, surveillance and counselling of individuals with high risk behaviours, as well as protection of the nation's blood supply. This paper examines the global response to the HIV pandemic and containment strategies appropriate to Singapore based on established epidemiological data.

A review of 58 patients in Singapore with significantly high anti-Toxoplasma serotitres.

Over a 42-month period, 58 patients presenting at the Singapore General Hospital were identified to have IgG anti-Toxoplasma serotitres of 1:1024 or higher by the indirect immunofluorescence test. This retrospective study examines the clinical presentation and management of these 58 patients. About 60% of these 58 patients had presented with lymphadenopathy, 20% with ocular symptoms and 14% with bad obstetric history. Almost all patients with IgG anti-Toxoplasma serotitres exceeding 1:1024 had presented with lymphadenopathy (with a modal titre of 1:4096). In contrast, most of the patients who presented with ocular symptoms or bad obstetric history had lower modal titres. The majority of patients who had presented with lymphadenopathy were Chinese, aged 21 to 35. Typically, they presented with a painless, mobile, solitary cervical node of three to four weeks duration as the only symptom, had biopsies where the histopathology was suggestive of toxoplasmosis, received no treatment and experienced no sequelae. The study concludes that: acute toxoplasmosis is common in Singapore and presents typically as asymptomatic cervical lymphadenopathy in a young Chinese; the incidence of congenital infection is also believed to be high, based on the number of cases presenting with fetal wastage and the prevalence of ocular toxoplasmosis; pigs in Singapore may constitute an important reservoir for the transmission of this disease to man.

Human immunodeficiency virus infection in Singapore--the first 50 cases.

As at 31 May 1990, fifty Singaporeans with the Human Immunodeficiency Virus (HIV) infection had been detected. Of these, nineteen had the Acquired Immunodeficiency Syndrome (AIDS). The majority of infected persons had been infected through sexual contact (homosexual 52%; bisexual 24%; heterosexual 20%) with men and women from countries where HIV infection was prevalent. The majority of infected patients (88%) were in the age range 20-39 years. There was one case of blood transfusion-associated AIDS. There were no infected paediatric or haemophiliac cases or intravenous drug use in any of the patients. A spectrum of AIDS-related opportunistic infections and cancers was observed, and Pneumocystis carinii pneumonia was the most frequent presentation. Thirteen patients with AIDS had died and the median survival time was about seven months.

Rapid in situ generation of DNA restriction endonuclease patterns for Neisseria gonorrhoeae.

Restriction endonuclease (RE) digestion patterns of 26 isolates of Neisseria gonorrhoeae representing different serovars of serogroups WI, WII, and WIII were generated by agarose pellet entrapment and in situ digestion with HinfI and BglII. The method was fast, simple, and reproducible, and stable RE patterns were produced from subsequent in vitro passages. The cost of culture materials was reduced considerably, and no toxic or flammable solvents needed to be used. Excellent resolution of DNA fragments of higher molecular weights was obtained as a result of minimal mechanical shearing of the DNA. The REs HinfI and BglII were discriminative in the fragment length ranges of 2 to 6.5 and 2.5 to 21.5 kilobases, respectively. On the basis of densitometric scanning of electrophoretograms generated by HinfI digestion, the 26 isolates representing 12 serovars were divided into seven groups. BglII was found to be more discriminative; 15 RE patterns were established among the 26 isolates. Patterns generated by both REs showed that there was no correlation between a particular RE pattern and a serovar, since strains with identical RE patterns were from different serovars. With the exception of two strains (D3 and D14), which demonstrated positive correlation when both enzymes were used, all strains with identical serovar patterns had different RE patterns.

The role of counterimmunoelectrophoresis in the identification of antibodies to extractable nuclear antigens.

Antibodies to saline-extractable nuclear antigens are hallmarks of autoimmune diseases including systemic lupus erythematosus (SLE), mixed connective tissue disease, progressive systemic sclerosis and Sjogren's syndrome. In our laboratory, we use counterimmunoelectrophoresis as a screening test and immunodiffusion as a confirmatory test to identify these autoantibodies. This study examines the drawbacks of such an approach. Though 17 out of 19 sera that formed ribonuclease sensitive lines with rabbit thymus extract on counterimmunoelectrophoresis were confirmed to have anti-RNP by immunodiffusion, sera of several different autoantibody specificities were seen to form ribonuclease resistant precipitin lines with the thymus extract on counterimmunoelectrophoresis. Having screened sera to have autoantibodies by counterimmunoelectrophoresis, the identity of some of these autoantibodies were not confirmed because of the poor sensitivity of immunodiffusion or because inappropriate controls had been used for the confirmatory immunodiffusion test. To check these drawbacks and to obviate the need for a confirmatory test, a modification of the current approach is suggested.

Evaluation of cerebrospinal fluid in asymptomatic late syphilis.

A prospective study of the cerebrospinal fluid (CSF) of 49 previously untreated patients with asymptomatic late syphilis of more than two year's duration was conducted. The sera of all patients had reactive fluorescent treponemal antibody absorption (FTA-Abs) tests or Treponema pallidum haemagglutination tests (TPHA), and reactive or non-reactive Venereal Disease Research Laboratory (VDRL) tests. Five (10%) patients had abnormal CSF findings; these included elevated protein in four of 49 and pleocytosis in three of 49. Asymptomatic neurosyphilis, diagnosed on the basis of a reactive VDRL test of the CSF, was present in one (2%) of 49 patients.

Enzyme immunoassay (Chlamydiazyme) for the detection of chlamydial antigen in endocervical specimens.

Endocervical swabs taken from 86 women were tested by the Chlamydiazyme enzyme immunoassay test to detect chlamydial antigens. Compared to cell culture, Chlamydiazyme was 100% sensitive and 90% specific. This test is suitable as alternative diagnostic method to cell culture and has the potential for automation.

Endocervical chlamydial infection in women with gonorrhoea.

Eighty six women attending Middle Road Hospital with endocervical gonococcal infection were evaluated. Chlamydia trachomatis was isolated in 27% of them. Women co-infected with C. trachomatis were similar to those with gonococcal infection alone in terms of demography, type of sexual contact, previous sexually transmitted disease, genitourinary symptoms, and clinical signs.

Relation of antinuclear fluorescence patterns and titres to autoantibody prevalence.

Different profiles of antinuclear antibodies (ANAs) have been found to be associated with different connective tissue diseases. Though several techniques have been developed to detect ANA, indirect antinuclear immunofluorescence (ANF) continues to be the most widely used and accepted. This study assesses the role of ANF as a screening test. The ANF patterns of 428 patients were studied. About 23% of patients with homogenous ANF patterns and 13% with speckled patterns were eventually shown to have raised anti-dsDNA titres. The percentage of patients harbouring raised titres of anti-dsDNA and the absolute titres were both seen to rise with the ANF titres. None of the patients with low ANF titres (1:40) harboured antibodies to extractable nuclea antigens (ENAs) whereas with increasing ANF titres the incidence of these autoantibodies was also seen to rise. Finally, it was noted that more than 80% of patients with high ANF titres demonstrated autoantibodies to ENA and/or dsDNA.

Chlamydial infection in female prostitutes in Singapore.

Three studies conducted in 1982, 1985 and 1988 investigated chlamydial infections in female prostitutes. In 1982, 115 prostitutes with culture-positive gonorrhoea were studied; 8% were coinfected with Chlamydia trachomatis. In 1985, 86 female prostitutes and in 1988, 100 female prostitutes attending for routine tests were examined. Chlamydia trachomatis was isolated in 12% and 9% of the cases, respectively. Gonorrhoea was detected in 10% and 11% of the cases. In the 1988 study, one (9%) of the 11 women with gonorrhoea had concomitant chlamydial infection. Syphilis was diagnosed in 3% of the female prostitutes investigated in 1988. Our findings indicate that concomitant chlamydial infection occurs in about 8-9% of female prostitutes with gonorrhoea and that 9-12% of the female prostitutes screened harboured Chlamydia trachomatis in their endocervices. A chlamydial control programme in prostitutes is desirable.

PIP: Studies conducted in 1982, 1985, and 1988 among female prostitutes in Singapore documented the need for a chlamydial control program in this population. In the 1982 study, which included 115 prostitutes infected with gonorrhea, Chlamydia trachomatis was recovered from 8%. In 1985 and 1988, the prevalence of endocervical chlamydia infection in prostitutes presenting for routine testing was 12% and 9%, respectively. The prevalence of gonorrhea in 1985 and 1988 was 10% and 11%, respectively. In the 1988 study, 1 of the 11 prostitutes with gonorrhea also harbored chlamydia. The prevalence of syphilis, ascertained only in the 1988 study, was 3%. These findings suggest that 8-9% of prostitutes with gonorrhea are co-infected with chlamydia. At present, prostitutes in Singapore are screened only for gonorrhea and syphilis. The high rate of C trachomatis infection recorded in this study indicates that chlamydia infection--on the increase in the West--should be added to this list. In addition, health education efforts are needed to stress the need for additional anti-chlamydial treatment. Currently utilized regimens for the treatment of gonorrhea are ineffective against C trachomatis.

Evaluation of an ELISA method for the measurement of antibodies to dsDNA.

Anti-dsDNA is found in 60-70% of patients with active, untreated systemic lupus erythematosus (SLE) and its detection serves as an important tool in the diagnosis and monitoring of these patients. This study evaluates the use of an Enzyme Linked Immunosorbent Assay (Elisa) to detect these antibodies. Its performance is also compared to the older, but established, method of detecting anti-dsDNA using Crithidia luciliae. The sera of the 56 normal healthy blood donors revealed a mean anti-dsDNA titre of 0.93mg% with a standard deviation of 0.23mg%. All 14 patients found to be negative by the Elisa method and 10 of the 11 patients found to have borderline anti-dsDNA Elisa titres were negative by immunofluorescence. 35 patients were found to harbour raised titres of anti-dsDNA by the Elisa method. All patients found to have anti-dsDNA titres exceeding 2mg% by the Elisa test were also positive by immunofluorescence. In fact, those with very high titres by the Elisa test were also strongly (titre greater than 1:100) positive by immunofluorescence. As a measurement of the kit's accuracy, the percentage of recovery of the activity of known amounts of antibody in a specimen fell within the range of about 89-104%. As a measurement of the kit's reproducibility, the coefficient of variation in the assayed titres of sample replicates was found to be 7.5% for within-batch assays and 9.7% for between-batch assays. The Elisa assay compared favourably to the immunofluorescence test in terms of enhanced sensitivity, quantitative approach with an objective end-point and the large number of samples that may be assayed simultaneously.