Anti-HIV-1 antibody testing using modified gelatin particle agglutination: a large field study.

Anti-HIV testing using gelatin particle agglutination (GPA) assay was investigated in parallel with ELISAs from routine service at Siriraj Hospital. In the first strategy, 174,032 sera from a patient population with an HIV-1 seroprevalence of 13.72 per cent were assayed using reduced volumes of GPA reagents, giving a cost reduction of 40 per cent. In the second strategy, 90,560 pregnant women and 48,936 emigrant workers with an HIV-1 seroprevalence of 2.2 per cent and 0.3 per cent, respectively, were tested in pools of 4 sera using the manufacturer's recommended volumes, giving a cost saving of 67 per cent. Overall, the sensitivity and specificity were almost identical with standard methods. Thus, parallel use of either modified GPA might be considered appropriate when testing large numbers of samples. However, both modified versions of GPA are not recommended as the first assay for diagnostic or blood bank screening especially in high prevalence of HIV infection.

Cytotoxic T lymphocyte responses to vaccinia virus antigens but not HIV-1 subtype E envelope protein seen in HIV-1 seronegative Thais.

The HIV-1 prime boost phase I/II vaccine trial using a recombinant canarypox vector, vCP1521, containing subtype E env (gp120), and subtype B env (gp41), gag and protease has started in Thailand. We have demonstrated that although 4 from 15 human immunodeficiency virus type 1 (HIV-1) seronegative Individuals showed cytotoxic T lymphocyte (CTL) responses to vaccinia virus antigens, none of them showed specific CTL responses to subtype E Env after in vitro stimulation. This preliminary study suggests that specific CTL responses to subtype E envelope detected in HIV-1 seronegative Individuals after vaccination should be considered as specific responses to the immunization.

Specific IgA antibody to Epstein-Barr viral capsid antigen: a better marker for screening nasopharyngeal carcinoma than EBV-DNA detection by polymerase chain reaction.

Nasopharyngeal carcinoma (NPC) is strongly associated with Epstein-Barr virus (EBV) infection. To assess whether EBV DNA detection by polymerase chain reaction (PCR) or presence of specific serum antibody to viral capsid antigen (VCA) was a better marker for screening NPC, nasopharyngeal tissues and blood samples from 58 NPC patients and 24 non-NPC patients (23 with laryngotracheal stenosis and 1 with chronic tonsillitis) were tested for the presence of EBV DNA and serum specific VCA antibodies, respectively. EBV DNA was detected in 56 (96.5%) of NPC patients and 15 (62.5%) of non-NPC controls, with predominantly EBV type A in both groups. On the other hand, specific VCA IgA antibody was detected in the majority of NPC patients: 52 (89.7%) while only 4 (16.7%) were detected in non-NPC controls. Therefore, specific VCA IgA antibody may serve as a better marker for screening NPC than EBV DNA detected by PCR.

Prevalence of type specific Epstein-Barr virus in the genital tract of genital herpes suspected patients.

A total of 62 clinical specimens from the genital tract of patients who were suspected of contracting genital herpes were investigated for HSV infection by the virus isolation method, and also investigated for the co-infection with EBV infection by detecting EBV DNA using nested PCR. HSV infection was diagnosed in 30 (48.4%) of the study cases, and so was EBV. EBV DNA was present in 17 (56.7%) of the 30 HSV positive samples. No correlation was found between the co-existence of these two viruses together. EBV DNA was detected in genital specimens of cervical, vaginal, urethral, and anal swabs. Ninety per cent of EBV belonged to type 1, and the remainder belonged to type 2 and mixed types. The role of EBV in genital tract infection needs to be further investigated.

Comparison between virus isolation method, Papanicolaou stain, immunoperoxidase stain and polymerase chain reaction in the diagnosis of genital herpes.

Papanicolaou (Pap) stain, immunoperoxidase (IP) stain and polymerase chain reaction (PCR) were evaluated against the virus isolation method for their sensitivity and specificity in the diagnosis of herpes simplex virus (HSV) infection in 96 women who were suspected of genital herpes. The result showed that the sensitivity of PCR, IP and Pap stain was 100, 92.0 and 62.7%, respectively, while the specificity was 76.2, 66.7 and 81.0%, respectively. PCR was even more sensitive than the virus isolation technique. As Pap stain is the technique routinely performed for diagnosing genital herpes in most of the hospitals in Thailand, its low sensitivity should be taken into consideration. Based on the investigation by all four techniques together, HSV infection was diagnosed in 91.6% of the cases suspected of genital herpes which reflected higher precision of the clinical diagnosis over Pap stain.

Prevalence of herpes simplex virus infection in patients suspected of genital herpes; and virus typing by type specific fluorescent monoclonal antibodies.

During the period between April 1994 and February 1996, a total of 154 female patients who attended the Clinic of Female Sexually Transmitted Diseases, Siriraj Hospital with clinical symptoms suspected of genital herpes were investigated for herpes simplex virus (HSV) infection by the virus isolation method in Vero cell cultures. Swabs from external genital lesions and the cervix from each patient were collected separately and used as the clinical specimens for isolation of HSV. The virus isolates were identified by indirect immunofluorescence (IIF) staining of the infected cell cultures using polyclonal HSV-2 specific antiserum which was reactive to common HSV antigens for both types of viruses. Typing of HSV was performed by direct IF using monoclonal antibody specific to HSV-1 or HSV-2. HSV was isolated from 78.6 per cent (121 of 154) of the cases studied; and among the infected cases, there were 47.9 per cent (58 of 121) in whom the infection involved both external genital lesions and cervixes, and 50.4 per cent (61) in whom the infection was limited to external genital lesions only. There were 2 cases (1.7%) in whom HSV was isolated from cervixes but not external genital lesions. Seventy-five HSV isolates were further subjected to typing. The present study showed that HSV-1 was accounted for 18.7 per cent (14 isolates), while HSV-2 took the remaining part of 81.3 per cent (61 isolates). The data demonstrated an increase in the prevalence of HSV-1 in genital herpes in our people.

Herpes simplex virus type-2, cytomegalovirus and Epstein-Barr virus infection in acute non A to E hepatitis Thai patients.

A significant number of acute non A to E hepatitis cases are reported in Thailand every year, and the etiologies of these cases are unknown. Members of the herpesviridae family have been reported to cause either a self limited or fatal hepatitis in a small proportion of patients in other parts of the world. To determine whether herpesviruses may play a role in acute non A to E hepatitis, sera from 32 acute hepatitis patients without markers for acute hepatitis A to E virus infection were examined for IgM to herpesvirus type 2 (HSV-2), cytomegalovirus (CMV), and Epstein-Barr virus (EBV) using commercially available assays. IgM to HSV-2 was detected in four sera, IgM to CMV was detected in one serum, and IgM to EBV was detected in one serum. All of the acute non A to E hepatitis patients recovered and none had underlying conditions associated with impaired immunity. These results suggest that herpesviruses should be considered in the differential diagnosis for Thai patients with hepatitis.

Shedding of HIV-1 subtype E in semen and cervico-vaginal fluid.

The uneven expansion of HIV-1 subtypes in each transmitted group raises the possibility that some viruses have less/more potential by qualitative/quantitative for heterosexual transmission compared to others. In Thailand, HIV-1 subtype E is mainly spread via heterosexual route and accounts for about 95 per cent of the infected cases. To determine whether high sexual infectivity of HIV-1 subtype E is due to the presence of a virus in genital fluid, we conducted a study to characterize shedding of HIV-1 in seminal and cervico-vaginal fluids of 30 HIV-1 subtype E infected Thai couples by PCR and virus isolation methods. All subjects had no HIV-associated diseases and other sexually transmitted diseases. HIV-1 subtype E DNA was detected in 22/30 (77.33%) of cervico-vaginal and also 22/30 (77.33%) of seminal fluid samples. The isolation rate of HIV-1 from semen and cervico-vaginal secretion was 36.67 per cent and 16.67 per cent, respectively. Number of HIV-1 subtype E DNA copies in the blood is reversely correlated with the number of blood CD4+ T cells, while that in genital fluid was not related to CD4+ T cell count. An increase in shedding of HIV- DNA subtype E in female genital tract compared to other HIV subtypes reported by other investigators might be one reason to explain the rapid spread of subtype E by heterosexual transmission in Thailand.

Immune status in congenital infections by TORCH agents in pregnant Thais.

A cross-sectional, sero-epidemiological survey of the prevalence of antibodies to TORCH agents during various stages of gestation revealed an overall rate of 13-15 percent having antibodies to Toxoplasma gondii; 85-87 percent, to rubella ; 79-81 percent, to herpes simplex virus (HSV); 100 percent, to cytomegalovirus (CMV); 82-86 percent, to human herpes virus type 6 (HHV-6); 1-2 percent, to hepatitis C virus (HCV). None of human T lymphotropic virus type I (HTLV-I) antibody was detected, and a prevalence of hepatitis B surface antigen (HBsAg) was 6 percent. Although a tendency was noted towards an increase of antibody detection to each TORCH agent as gestation progressed, a statistically significant increase in antibodies titer and specific IgM antibody was found with regard to CMV. These results suggest an increase in CMV infection or reactivation during pregnancy whereas an increase in the other TORCH infections was not obvious.

V3 peptide enzyme immunoassay for serotyping HIV-1 infected pregnant Thais.

Previous molecular epidemiological studies show that at least 2 subtypes of HIV-1 circulate in Thailand. HIV-1 subtype B or Thai genotype B was associated with an early epidemic and was prevalent in intravenous drug users. Meanwhile, HIV-1 subtype E or Thai genotype A was becoming widespread among heterosexuals. We studied the HIV subtypes of 161 HIV-1 seropositive pregnant women. Of these, 143 pregnant patients (88.8%) tested positive for subtype E alone and 8 women (5.0%) had evidence of infection with subtype B alone. There was serologic evidence of infection with a mixture of subtypes in 7 women while the infecting subtype could not be identified in the remaining 3 women. This result agrees with previous information that subtype E predominates in Thai heterosexuals.

Serological response to respiratory syncytial virus infection in pediatric patients with a comparison to immunofluorescence and virus isolation.

The serological response to respiratory syncytial virus (RSV) in 125 pediatric patients hospitalized with acute lower respiratory infection was investigated by enzyme linked immunosorbent assay (ELISA) for specific immunoglobulin (Ig) A, IgG, and IgM and complement fixation (CF) test. By ELISA, a 4-fold rise in IgG titre in paired sera was most commonly found, followed by a rise in IgA and IgM titres. Investigation by ELISA and CF leads to the suggestion that major CF activity against RSV antigens resides in the IgG and not the IgA and IgM classes. No case with CF activity failed to be diagnosed by ELISA. The youngest infant who could develop seroconversion was one month old, nevertheless two children older than two years could not. When the three diagnostic methods were compared, ELISA serology was the most sensitive followed by indirect immunofluorescence (IIF) for antigen detection and virus isolation, respectively, ELISA could diagnose RSV infection in 45% of the study cases, whereas IIF and virus isolation only diagnosed 26% and 14%, respectively. Half of the cases was diagnosed by all of the three methods together.

Incidence of Mycoplasma pneumoniae, Chlamydia trachomatis, and viral infections in pneumonia cases under six months of age, Bangkok, Thailand.

The incidence of infections by Mycoplasma pneumoniae, Chlamydia trachomatis and respiratory viruses was investigated in 76 pneumonic patients aged under 6 months who attended Ramathibodi and Siriraj Hospitals in Bangkok during two study periods. M. pneumoniae infection was not found in any case from either hospital by serological diagnosis. By the isolation method, C. trachomatis infection was found in 7(16.7%) of 42 patients from Ramathibodi Hospital and 5(21.7%) of 23 patients from Siriraj Hospital with the average male:female ratio of 2.6:1; and 91.7% of the infected cases were under 3 months old. Laboratory diagnosis of respiratory virus infection was performed by indirect immunofluorescence (IIF), isolation, and by antibody detection. Data from Ramathibodi Hospital showed that 11 (24.4%), 4 (8.9%), 3 (6.7%) of the 45 patients were infected by respiratory syncytial virus (RSV), adenoviruses, parainfluenza virus type 3, and some other viruses, respectively; infection rates of 10 (32.3%), 4 (12.9%), 1 (3.2%) and 1 (3.2%) by those viruses respectively, were observed in the 31 patients from Siriraj Hospital.

The abbreviated 2-1-1 schedule of purified chick embryo cell rabies vaccination for rabies postexposure treatment.

During August 1988 to January 1990, the immunogenicity and safety of purified chick embryo cell rabies vaccine (PCEC) given by the conventional and abbreviated regimens in 82 vaccinees moderately to severely exposed to laboratory proven rabid animals were studied. The 16 vaccinees received PCEC six doses as conventional schedule on days 0, 3, 7, 14, 28 and 90, the 11 vaccinees received six doses of PCEC plus human rabies immune globulin (HRIG) on day 0. The 29 vaccinees received an abbreviated schedule of PCEC as two doses on day 0, one dose each on days 7 and 21 and the 26 cases received PCEC abbreviated schedule plus HRIG on day 0. The kinetics of the neutralizing antibodies on days 0, 7, 14, 28, 56, 180 and 365 were studied for comparative purpose. All vaccinees had high antibody levels from day 14 which last longer than a year and were safe after one year follow up. The adverse reactions of the vaccine were mild and self-limited.

Detection of Epstein-Barr virus DNA and HHV-6 DNA in tissue biopsies from patients with nasopharyngeal carcinoma by polymerase chain reaction.

A total of 34 tissue biopsies were collected from nasopharyngeal carcinoma (NPC) patients and 5 controls with non-NPC. Extracted DNA from tissue biopsies were analyzed for presence of specific gene sequences to EBV type A and type B, and HHV-6 by polymerase chain reaction (PCR). The different sequences of EBV type A and B were parts from the highly divergent forms of the EBV nuclear antigen 2 (EBNA 2). The PCR amplified products for EBNA 2A and EBNA 2B were 115 and 119 base pairs respectively whereas that of HHV-6 DNA was 776 base pairs. The results demonstrated that EBV DNA was detected in 32 of 34 cases (94.1%): 28 (82.3%) with type A, 2 (5.9%) with type B, and 2 (5.9%) with both types. EBV DNA of type A could be detected 1 (20%) of 5 controls. HHV-6 DNA was in 5 of 34 samples (14.7%) whereas HHV-6 DNA was not detectable in biopsy tissues from controls. The results show that in the NPC patient group, A type of EBV is predominant. Detection of HHV-6 DNA in patients group only might be resulted from reactivation of a latent infection or association with EBV-induction of NPC.

Prevalence of human herpesvirus 6 in Thai people and its correlation with human immunodeficiency virus type 1 infection.

Human herpesvirus 6 (HHV-6) is a human herpesvirus isolated from patients with various lymphoproliferative disorders and acquired immunodeficiency syndrome (AIDS). The prevalence of HHV-6 infection and its correlation as a cofactor in pathogenicity of HIV infection was investigated in serum samples from 365 healthy volunteers at various age groups, 50 persons at risk for HIV-1 infection, and 90 HIV-1 seropositive individuals. Sera were screened and titrated for antibodies against HHV-6 by a standard indirect immunofluorescence assay on an acetone fixed HHV-6 infected HSB2 cells. The data show high prevalence of HHV-6 in Thailand (71.7%) and the infection is acquired early in life. Prevalence of anti-HHV-6 IgG antibodies was not strikingly different among people at risk for HIV infection, asymptomatic HIV-1 infected cases, and aged-matched controls with low risk for HIV-1 infection. The AIDS cases showed high titers of anti-HHV-6 IgG antibody and high rates for presence of anti-HHV-6 IgM antibody (33.3%) which suggests higher prevalence of HHV-6 infection by either reactivation of an earlier HHV-6 infection or a new primary infection.

Prevalence of IgA specific antibodies to Epstein-Barr virus capsid and early antigens in nasopharyngeal carcinoma.

Ninety-one patients with nasopharyngeal carcinoma (NPC), and 164 age-matched healthy controls were tested for presence of IgA antibodies to Epstein-Barr virus capsid antigen (VCA) and early antigen (EA) in their sera by indirect ELISA using "EBViral DETECT" commercial test kit. IgA anti-VCA was found in 76 (83.5%) of NPC patients and 16 (9.8%) of the controls. Meanwhile, IgA anti-EA was found in 72 (79.1%) of NPC patients and 21 (12.8%) of the controls. In a parallel study by indirect immunofluorescence test (IIF), IgA anti-VCA was found in 77 of 91 (84.6%) NPC patients and 22 of 142 (15.5%) controls. The prevalence rates of anti-VCA as screened by ELISA and IIF were very similar suggesting that neither one of the two tests can be used alternatively depending on the purpose and facilities in each individual laboratory. IgA antibodies to VCA and EA were more prevalence in NPC patients than those in the controls, the finding which again supported the association between EBV and NPC as was suggested in many other reports.

Epstein-Barr virus serological markers for nasopharyngeal carcinoma in Thailand.

The present study reports on the prevalence of specific IgA and IgG antibodies to EBV viral capsid antigen in nasopharyngeal carcinoma (NPC) patients with different histological types of carcinoma and their age-matched controls by the indirect immunofluorescence test, using the B-95-8 lymphoblastoid cell line as source of viral capsid antigen. EBV specific IgG was found in almost all the study cases, and antibody titers were significantly higher in the NPC patients than in non-cancer controls. GMT of anti-EBV IgG in NPC patients, patients with other malignant diseases, and those with non-malignant diseases were 371.5, 97.7 and 35.5, respectively. Anti-EBV specific IgA was more specific to NPC than was IgG, and was present in 86.5% (83 of 96) cases of NPC patients, 6.6% (2 of 30) of patients with other cancers, and 3.1% (3 of 97) cases of non-malignant diseases. A weak correlation between level of anti-EBV IgA in NPC patients was observed (r = 0.3). EBV IgA was found in all histological types of NPC, ie, WHO types 1, 2 and 3, but WHO type 1 was rare among NPC patients in Thailand. Use of anti-EBV IgA for monitoring cancer therapy is to be further investigated.

Rabies and post-exposure prophylaxis in Thai children.

Thailand is an endemic area for rabies, with approximately 300 human deaths reported annually. More than half of the rabies patients are children under 14 years of age. This paper reports clinical data of paediatric rabies cases occurring from 1980 to 1986, and the protective efficacies of human diploid cell rabies vaccine (HDCV) and purified Vero cell rabies vaccine (PVRV) in children exposed to rabid animals. The analysis of 120 medical records revealed that rabies in children had incubation periods which ranged from less than fifteen days to more than three months, but generally between one to three months. The most frequent symptoms observed in the patients were hydrophobia, restlessness, fever, vomiting and aerophobia. Most of the rabid children admitted to hospital died within 24 hours. HDCV was administered to 50 children exposed to rabies with the cumulative dosages of 327 ml. All patients survived without serious adverse effects during a-two year follow-up. Mild reactions were seen in 1.5 percent (5/327 doses). Unfortunately, levels of rabies antibody in these vaccinees were not determined. Among another series of children exposed to rabid animals, comprising 27 individuals who received a total of 168 doses of PVRV, only mild local reactions were seen in 6 subjects. No rabies deaths were reported in 2 years of follow-up. The children who received PVRV either with or without human rabies immune globulin developed similar levels of rabies neutralizing (NT) antibody, which reached the high titers on day 30. At one year after the first dose of vaccination, all vaccinees still had NT antibody at titers higher than 0.5 IU/ml.(ABSTRACT TRUNCATED AT 250 WORDS)