AIDSVAX (MN) in Bangkok injecting drug users: a report on safety and immunogenicity, including macrophage-tropic virus neutralization.

A randomized, double-blind, placebo-controlled phase I/II study of AIDSVAX (MN) was conducted among injecting drug users in Bangkok, Thailand. Four doses of vaccine (300 microg of MN-rgp120 in alum) or placebo (alum) were given at study entry and at 1, 6, and 12 months. The objectives of the study were to evaluate (1) the feasibility of conducting vaccine trials in this population; (2) the safety of this candidate AIDS vaccine; and (3) the immunogenicity of this vaccine. Thirty-three volunteers (22 vaccine and 11 placebo recipients) were recruited. None were lost to follow-up during the 18-month study. Mild reactogenicity was noted, which was similar in both vaccine and placebo recipients. The vaccine induced anti-HIV-1 antibody in all vaccine recipients. Maximal titers of binding antibodies of MN-rgp120 and the V3 domain of MN-rgp120 were induced after the third (6 month) dose while maximal neutralizing antibodies followed the fourth (12 month) dose. The vaccine-induced antibodies from several volunteers were capable of neutralizing macrophage-tropic, subtype B viruses (301660 and JRCSF) detected in a PBMC-based assay. Binding and neutralizing antibodies declined about 10-fold in the 6 months after the last boost. Two vaccinees became infected during the trial, both with subtype E viruses. A phase III efficacy trial, using a bivalent gp120 vaccine containing antigens from a subtype B virus (MN) and a subtype E virus (A244), was initiated in March 1999 in injecting drug users in Bangkok.

Diagnosis of Penicillium marneffei infection by quantitation of urinary antigen by using an enzyme immunoassay.

Penicillium marneffei is a major cause of opportunistic infection in patients with AIDS in north and northeastern Thailand. A method for the quantitation of P. marneffei antigen in urine was developed by using fluorescein isothiocyanate-labelled purified rabbit hyperimmune immunoglobulin G in an enzyme-linked immunosorbent assay. This method was evaluated with 33 patients with culture-proven penicilliosis and 300 controls (52 healthy subjects, 248 hospitalized patients without penicilliosis) from the same area in which penicilliosis is endemic. Urinary antigen was found in all 33 (100%) patients with penicilliosis, with a median titer of 1:20,480. With undiluted samples, 67 (27%) of 248 hospital patients and 3 (6%) of 52 healthy controls were reactive. At a cutoff titer of 1:40, the urine antigen detection assay had a diagnostic sensitivity of 97% and specificity of 98% (positive predictive value, 84%; negative predictive value, 99.7%). This test offers a valuable and rapid method for the diagnosis of penicilliosis in patients with AIDS and could be a useful addition to conventional diagnostic methods in areas in which penicilliosis is endemic.

Changes of liver functions after albendazole treatment in human gnathostomiasis.

Ninety-eight out-patients of the Hospital for Tropical Diseases, Bangkok with clinical diagnosis of cutaneous gnathostomiasis were studied. All patients were treated with albendazole at a dosage of 400 mg (two tablets) twice daily for 14 days. They were seen periodically on day 0, day 14, day 28, day 195 and 1 year after treatment with laboratory investigations for any side effects of the treatment. There was a statistically significant increase of total protein, albumin, alkaline phosphatase (ALP), aspartate aminotransferase (AST) and alanine aminotransferase (ALT) values when comparing the different periods. The abnormal results are clearly indicated in AST and ALT values (liver enzyme) especially on day 14 both male and female patients had highest levels. No significant association with time was found in ALP value.

Endogenous interleukin-10 modulates proinflammatory response in Plasmodium falciparum malaria.

Tumor necrosis factor-alpha (TNF-alpha), interleukin (IL)-1beta, and IL-6 are implicated in the pathogenesis of severe Plasmodium falciparum malaria. In this study, the effect of IL-10 on their production by peripheral blood mononuclear cells (PBMC) from acutely infected patients was examined. Exogenous IL-10 inhibited malarial antigen-induced cytokine production by reducing mRNA accumulation. Maximal inhibition occurred when IL-10 was added in the first 2 h of stimulation. Conversely, the addition of anti-IL-10 markedly enhanced TNF-alpha, IL-1beta, and IL-6 production. The effect was significantly greater on PBMC from patients with uncomplicated infection than PBMC from patients with severe disease. Kinetics studies showed that TNF-alpha, IL-6, and IL-1beta were produced within 2-4 h of stimulation, while IL-10 was first detectable after 8 h. These findings suggest that IL-10 counter-regulates the proinflammatory response to P. falciparum. Severe falciparum malaria may be associated with an inadequate negative feedback response by IL-10.

SPf66 malaria vaccine is safe and immunogenic in malaria naive adults in Thailand.

In preparation for an efficacy trial of malaria vaccine SPf66 in Thailand, a series of overlapping Phase I trials were conducted of US-manufactured SPf66. Here, two clinical lots were evaluated for safety and immunogenicity in a combined open-label trial. Eleven healthy, malaria naive, 18-44 year-old Thai men and women received three doses by subcutaneous injection in alternate arms at 0, 1 and 6 months. Safety was assessed by monitoring local and systemic reactogenicity and laboratory parameters. Common side effects were mild erythema, induration and tenderness at the site of injection which resolved within 24-48 h. At third immunization, two volunteers developed acute bilateral reactions with induration, erythema and pruritus limited to the sites of the second and third immunizations. Eight of 11 volunteers sero-converted by ELISA, six of whom would be classified as high responders by Colombian standards. Eight of 11 volunteers developed a lymphoproliferative response to the SPf66 antigen. Side effects were more common and antibody and lymphoproliferative responses greatest, among the four female volunteers. This initial study of SPf66 malaria vaccine in Asia constitutes an essential link between the initial Phase I study in the US and subsequent field studies in a semi-immune population in a malaria endemic area of Thailand. This study further establishes comparability of US-manufactured SPf66 with that of Colombian provenance and substantiates the validity of the subsequent negative efficacy results of SPf66 in a field trial in Thailand.

Semi-quantitative measurement of Plasmodium falciparum antigen PfHRP2 in blood and plasma.

Plasmodium falciparum histidine rich protein 2 (PfHRP2) antigen was measured semi-quantitatively in whole blood, plasma, and supernatants and red blood cells of cultures in vitro using the dipstick ParaSight-F test and also by a quantitative antigen-capture enzyme-linked immunosorbent assay (ELISA). In vitro, PfHRP2 was secreted mainly during the second half of the asexual cycle with a marked rise during schizont development and rupture. The total PfHRP2 secreted before schizogony corresponded to approximately 4% of that contained in the red blood cells. In samples from 55 patients with acute falciparum malaria, the level of detection by ELISA corresponded to parasitaemias of 100/microL for whole blood and 1600/microL for separated plasma. Whole blood PfHRP2 levels were correlated significantly with admission parasitaemia (r = 0.76, P < 0.0001) and the stage of parasite development (r = 0.43, P < 0.01). Although whole blood PfHRP2 concentrations were higher in severe malaria, plasma concentrations of PfHRP2 were considerably higher in severe malaria (median titre 1:320, range zero to 1:1280) than in uncomplicated malaria (median titre 1:5, range zero to 1:80; P < 0.0001). The ratio of whole blood to plasma PfHRP2 was lower in severe than in uncomplicated malaria (median 4, range 0.25 to 256, versus 64, range 4 to 1280; P < 0.0001). With plasma samples the intensity of colour change on the dipstick correlated well with more precise measurement of optical density in the ELISA (r = 0.88, P < 0.0001). These results suggest that measurement of PfHRP2 in plasma could provide an alternative approach to the assessment of the parasite biomass, and thus prognosis, in severe malaria, and that this could be done simply by using the currently available dipsticks.

Risk behaviours and comprehension among intravenous drug users volunteered for HIV vaccine trial.

Out of 91 volunteers enrolled for the HIV vaccine trial, only 33 volunteers were eligible for vaccination. Of 33 volunteers recruited, 59 per cent of them had incomes of more than 5,000 Baht/ month. The median duration of drug addicts was 15 years (range 1-26 years) and 42 per cent never used condoms during sexual intercourse. As far as consent comprehension was concerned, all of them understood.

PIP: With HIV seroprevalence rates of 35-40%, Thai intravenous drug users are an obvious target group for anti-HIV vaccine trials. The Ministry of Public Health of Thailand recently approved clinical trials of the Genentech MN rgp 120/HIV-1 candidate vaccine among recovering intravenous drug users enrolled in a methadone treatment center in Bangkok. To evaluate safety and immunogenicity factors and assemble a relatively low-risk group, 91 potential volunteers from this center were screened (history, physical examination, and blood and urine analysis). Only 33 passed all of the screening procedures; the major causes of ineligibility were HIV seropositivity (14 cases) and impaired liver function test (10 cases). The 33 eligibles had a median duration of drug addiction of 15 years. The majority had a secondary education or above and a monthly income exceeding 5000 Baht (US$200). Never-use of condoms was acknowledged by 14 of the eligible volunteers, but all 33 denied relationships with commercial sex workers. Among those selected for the trial, 23 believed a successful vaccine against AIDS is likely and 30 thought the vaccine they received was effective; however, 12 were concerned the vaccine would enhance disease progression. After 12 months of counseling and participation in the vaccine trial, nine volunteers were drug and methadone-free. Overall, these findings indicate that intravenous drug users can successfully participate in vaccine trials if there is adequate pre-screening.

Interleukin-10 inhibits tumor necrosis factor production but not antigen-specific lymphoproliferation in acute Plasmodium falciparum malaria.

In vivo interleukin (IL)-2, IL-4, IL-10, and interferon (IFN)-gamma production was measured at the mRNA transcript and protein levels in patients acutely infected with Plasmodium falciparum and during convalescence. Both IL-10 and IFN-gamma but not IL-2 were produced regardless of the patients' clinical severity. IL-4 production was variable. Circulating IFN-gamma and IL-10 were significantly higher in patients with severe disease (P < .01 and .001, respectively). In vitro stimulation of peripheral blood mononuclear cells (PBMC) by malarial antigens during acute infection showed that although there was no lymphoproliferation, the cells could produce IL-10 and IFN-gamma. Recombinant human IL-10 completely abolished in vitro tumor necrosis factor (TNF)-alpha production in response to malarial antigens, as well as the antigen-specific proliferative response of convalescent patients. However, anti-IL-10 was insufficient to restore proliferation of PBMC from acutely infected patients. These findings suggest that IL-10 may have an important negative feedback action on the production of inflammatory cytokines in acute falciparum malaria without contributing to the defect in antigen-specific proliferation.

Detection of Pseudomonas pseudomallei antigen in urine for the diagnosis of melioidosis.

An enzyme-linked immunosorbent assay using a fluorescein isothiocyanate (FITC)-anti-FITC amplification system, has been developed to detect Pseudomonas pseudomallei antigen in urine. The assay was evaluated in 135 patients with acute melioidosis, 194 hospitalized patients with other disorders, and 40 healthy controls. Antigen was detected in the urine of 123 (91%) patients with melioidosis. Urinary antigen was found in 85 (96%) of 89 patients with septicemic melioidosis, all six patients with P. pseudomallei urinary tract infection, and 32 (80%) of 40 patients with other localized infections. Antigen was not detected in the urine of 40 healthy individuals, but the urine of 16 (8%) of 194 hospitalized patients with diagnoses other than melioidosis gave a positive result. Of the false-positive results, 13 of 16 were associated with bacteriuria > or = 10(4) colony-forming units/ml. At a cutoff titer of 1:10, the sensitivity and specificity of the test were 81% and 96%, respectively. Enzyme immunoassay detection of urinary antigen is a valuable and rapid laboratory test for the early diagnosis of acute melioidosis.

A simplified and economical intradermal regimen of purified chick embryo cell rabies vaccine for postexposure prophylaxis.

Healthy volunteers were randomized to receive either intradermal purified chick embryo cell rabies vaccine (PCEC) alone (0.1 ml at each of two sites on days 0, 3 and 7, and at one site on days 28 and 90) (n = 81), or intradermal PCEC with one dose of human rabies immunoglobulin (HRIG) intramuscularly at 20 IU kg-1 on day 0 (n = 52). Neutralizing antibody (NAB) was detectable in every volunteer, in both groups, from day 14 up to day 365. The peak NAB occurred on day 28 in both groups. No significant suppressive effects of HRIG on NAB response were observed. Side-effects were mild and self-limiting. These preliminary results suggest that this simplified low-dose intradermal regimen could be an alternative schedule in rabies postexposure prophylaxis, resulting in lower overall costs.

Malaria in splenectomized patients: report of four cases and review.

The spleen plays a central role in host defense against malaria in animals. Its role in human malaria is less well established. The spleen may contribute to protection against human malaria by mediating humoral or cellular immune responses or by clearing both rheologically and immunologically altered host erythrocytes. This report describes Plasmodium falciparum and Plasmodium vivax infections that occurred after splenectomy in one nonimmune and three partially immune Thai adults. The clinical course was uncomplicated for all four patients, and parasite clearance was delayed only in the nonimmune patient. In three patients with falciparum malaria, humoral and cellular immune responses to blood-stage antigens during the acute infection and convalescence were similar to those of individuals whose spleens were intact. These findings suggest that the spleen may not be essential for the processes leading to parasite clearance in partially immune, splenectomized patients. Further studies on the course of malarial infections in splenectomized patients are crucial for clarifying the role of the spleen in host defense against human malaria.

Pasteur oral cholera vaccine: studies of reactogenicity, clinical acceptability and immunogenicity in human volunteers.

Pasteur cholera vaccine consists of isolated antigenic fractions from V. cholerae El Tor Ogawa and Inaba. Enteric coated microgranules were prepared from antigen lyophilisate. Three doses of this vaccine were administered orally to 19 healthy young Thai adults at one week intervals. None of the volunteers experienced untowards reactions. The vibriocidal antibody responses manifested a significant antibody rise (> or = 4 fold) to serovar Inaba in 8 vaccinees (42.1%) and Ogawa in 4 (21.1%). Five and 6 vaccinees (26.3% and 31.6%) showed a > or = 4 fold rise of IgG and IgA anti-LPS, respectively.

Albendazole stimulates outward migration of Gnathostoma spinigerum to the dermis in man.

Human gnathostomiasis is characterized by space-occupying inflammatory lesions and/or hemorrhage as a result of the migration of, very often, a single larva of Gnathostoma spinigerum. Intermittent cutaneous migratory swellings occurring over years is the most common manifestation and the rare cerebral invasion may be fatal. There are currently no effective anthelminthics for this infection. During a double-blind randomized placebo control trial evaluating the efficacy of albendazole in cutaneous gnathostomiasis at a dosage of 400 mg twice daily for two weeks, it was observed that gnathostome larvae tended to migrate outward as a result of the treatment so that they could be recovered by excisional biopsy or by picking with a needle. In the placebo-treated group (N = 40), no such migration was observed during the 8,470 patient-days of follow-up while in the albendazole-treated group (N = 41) there was one worm in an excisional biopsy done on day 16 and two worms were removed from the skin by the patients themselves on days 8 and 0. Assuming that the period of drug exposure of the gnathostomes was the 14 days of albendazole administration plus another washout period of 7 days (equivalent to 20 half-lives of the active detectable metabolite), the total patient-days of albendazole exposure was 830. The rate of outward migration of gnathostomes in the drug treated group (3 per 830 patient-days) was significantly (p < 0.0001) higher than in the placebo group (0 per 8,470 patient-days).(ABSTRACT TRUNCATED AT 250 WORDS)

Cytoadherence and ultrastructure of Plasmodium falciparum-infected erythrocytes from a splenectomized patient.

In malarial infections of primates, the spleen has been shown to modulate parasite antigen expression on the surfaces of infected erythrocytes. The processes affected include cytoadherence, which is central to the pathophysiology of severe falciparum malaria, and the related phenomenon of rosette formation. In this study, the cytoadherence and rosette formation behaviors of Plasmodium falciparum-infected erythrocytes from a splenectomized patient were examined during the first erythrocytic cycle in vitro. Ultrastructural studies were also performed. Infected erythrocytes were found to cytoadhere to C32 melanoma cells via leukocyte differentiation antigen CD36 but not intercellular adhesion molecule 1. They also displayed on their surfaces electron-dense knobs similar in structure and density to those on infected erythrocytes from intact hosts. These findings may reflect a stable cytoadherent phenotype of the parasite isolate that is unaffected by the absence of the spleen. Alternatively, the modulating role of the spleen may have been assumed by other organs of the mononuclear phagocytic system in a previously infected individual. No rosette formation was observed, but as not all natural isolates form rosettes, this observation may or may not be related to the asplenic status of the patient. Parasite and host factors appear to be important in determining the effect of splenectomy on cytoadherence and rosette formation in human falciparum malaria.

Randomised trial of artesunate and mefloquine alone and in sequence for acute uncomplicated falciparum malaria.

The increasing frequency of therapeutic failures in falciparum malaria in Thailand shows an urgent need for effective drugs or drug combinations. Artesunate, a qinghaosu derivative, is effective in clearing parasitaemia rapidly, but the recrudescence rate can be as high as 50%. We have compared artesunate followed by mefloquine with each drug alone in acute, uncomplicated falciparum malaria. 127 patients were randomly assigned treatment with artesunate (600 mg over 5 days), mefloquine (750 mg then 500 mg 6 h later), or artesunate followed by mefloquine. All patients were admitted to hospital for 28 days to exclude reinfection. Cure was defined as no recrudescence during the 28 days' follow-up. The cure rates for mefloquine and artesunate alone were 81% (30/37 patients) and 88% (35/40); the combination was effective in all of 39 patients. Fever and parasite clearance times were significantly shorter in the groups that received artesunate than in the mefloquine-only group. The frequency of nausea and vomiting was slightly, but not significantly, higher among patients who received both drugs than in the other groups. The combination of artesunate followed by mefloquine is highly effective and well tolerated in patients with acute, uncomplicated falciparum malaria in Thailand.

PIP: Physicians enrolled 127 patients who were admitted to the Bangkok Hospital for Tropical Diseases in Thailand with acute, uncomplicated falciparum malaria between January-May 1991 into a randomized clinical trial of 3 oral treatments: artesunate, mefloquine, and artesunate followed by mefloquine. At the end of 28 days, 88% of patients who received only artesunate (total dose 600 mg), 81% of those who received only mefloquine (total dose 1250 mg), and all patients who received both artesunate and mefloquine were cured. Artesunate reduced the parasite count by 90% within 24 hours of 1st treatment. The 2 groups that received artesunate experienced considerably more rapid reduction in parasitemia and in fever than the group that received only mefloquine (p.002). Mefloquine was more adept than artesunate at clearing residual parasites. Mefloquine-treated patients experienced slightly more headaches and dizziness while they still had malaria than the other 2 groups. The physicians believed that these symptoms could actually have been due to the acute malaria infection. Patients who were on the sequential artesunate-mefloquine regime experienced more nausea and vomiting than the other groups, but the differences were insignificant. The combination therapy with artesunate and mefloquine was very effective and patients with acute, uncomplicated malaria tolerated it well.

Clinical and bacteriological studies of El Tor cholera after ingestion of known inocula in Thai volunteers.

Twenty-six healthy adult Thai volunteers were recruited for clinical and bacteriological studies of cholera induced by oral inoculation with Vibrio cholerae El Tor Inaba strain N16961. Vibrio dosages of 0.3 x 10(4), 1.6 x 10(5) and 1.9 x 10(6) c.f.u. were given to three groups of five volunteers, and 2.0 x 10(7) c.f.u. to 11 volunteers. Diarrhoeal attack rates correlated positively with the size of the inocula (p less than 0.01). It was estimated that a diarrhoeal attack rate of 90% (ED90) would be achievable by inoculation of 1.3 x 10(7) c.f.u. of the organisms. There were no significant differences between the groups in the latent period to positive stool culture, maximum vibrio count per gram of stool and duration of stool positivity. The ED90 of V. cholerae obtained may be used as a challenge dose in subsequent studies on protective efficacy of cholera vaccines in Thai adult volunteers.

Health status of Thai volunteers in a cholera vaccine trial.

One hundred and seventy-one male adults were screened in recruitment of volunteers for a cholera vaccine trial. A full medical history and a physical examination were performed on each subject. The percentages of subjects vaccinated against cholera and typhoid within twelve months were 4 and 1 per cent respectively, while 88 and 15 per cent respectively had been vaccinated more than a year. Biochemical screening revealed abnormal liver function tests in 40.7 per cent, specifically alkaline phosphatase (8%), glutamic oxaloacetic transaminase (8%), glutamic pyruvic transaminase (4.7%), total bilirubin (10%) and globulin (34%). Ten (6%) of the volunteers were positive for hepatitis B surface antigen (HBs-Ag). The total white cell count was elevated in 13.5 and 81.9 per cent had eosinophilia. Stool examination revealed infection with Hookworm (54.9%), Opisthorchis viverrini (29.8%), Strongyloides stercoralis (5.3%), Endolimax nana (3.5%), Giardia lamblia (5.3%) and Taenia saginata (2.9%). Few volunteers (13.4%) had abnormal microscopic examination of urine sediment. Only 57 subjects were considered suitable to be volunteers. Each of these subjects had no significant past medical, surgical or psychological illness. None had been vaccinated against cholera within the previous 12 months and no subject had abnormalities on physical examination or routine biochemical and haematological screening. The large number of subjects excluded from recruitment (67%) emphasized the importance of proper screening of volunteers for any vaccine trial.

Relationship of vibriocidal antibodies to history of cholera vaccination in Thai adult volunteers.

Vibriocidal antibodies were determined by microtechnique in 5 groups of Thai adult volunteers who had never received or had received cholera vaccination within one year, more than one to five years ago, more than five to ten years ago and more than ten years ago respectively. Detailed questionnaires about socioeconomic status, educational levels and environmental factors were presented to every volunteer. There were no differences statistically in incomes, educational levels and environmental factors among the groups. It was found that the reciprocal geometric mean titers of antibodies in volunteers who had never received cholera vaccination was generally low. The reciprocal geometric mean titers of the volunteers who had received cholera vaccination within one year were statistically different from other groups (p = 0.05). There was no correlation between blood groups of volunteers and vibriocidal antibodies.

Preliminary assessment of the safety and immunogenicity of live oral cholera vaccine strain CVD 103-HgR in healthy Thai adults.

A single dose (5 x 10(8) organisms) of attenuated A- B+ Vibrio cholerae classical Inaba recombinant vaccine strain CVD 103-HgR or placebo was administered to 24 healthy young Thai adults in a randomized, placebo-controlled, double-blind trial of safety and immunogenicity. None of the volunteers experienced untoward reactions. The vaccine strain was recovered from 2 of 12 vaccines. The vibriocidal antibody response (the best immunological correlate of protection) was good: 11 of 12 vaccinees (92%) manifested significant serotype-homologous Inaba antibody rises with a peak reciprocal geometric mean titer (RGMT) postvaccination of 3,417; 9 of 12 exhibited significant serotype-heterologous Ogawa antibody rises (prevaccination RGMT, 180; peak RGMT, 2,874). Nine of 12 vaccinees had significant rises in serum antitoxin. None of the controls exhibited rises in vibriocidal or antitoxic antibody. This preliminary study further confirms the safety and immunogenicity of CVD 103-HgR live oral cholera vaccine and paves the way for larger community studies of this candidate cholera vaccine.

Failure of interferon alfa and tribavirin in rabies encephalitis.

OBJECTIVE: To test the effect of interferon alfa and tribavirin (ribavirin) in patients with rabies encephalitis. DESIGN: An open trial of chemotherapy and intensive care in patients with early rabies. SETTING: The intensive care unit of a Bangkok hospital. PATIENTS: Four conscious men with clinical rabies encephalitis. INTERVENTIONS: Rapid virological diagnosis of rabies. Treatment with intravenous and intraventricular injections of high doses of lymphoblastoid interferon alfa in three patients and tribavirin in one patient. Intensive care was given throughout. MAIN OUTCOME MEASURES: Rabies infection confirmed by antigen detection and virus isolation. Rabies neutralising antibody and specific IgM sought in serum and cerebrospinal fluid. Interferon concentrations monitored before and during treatment in three patients. RESULTS: Interferon alfa treatment produced high concentrations in serum and cerebrospinal fluid. All four patients died after 5 1/2 to 12 1/2 days of treatment with no evidence of virostatic or clinically beneficial effects from either treatment. CONCLUSION: Interferon alfa treatment is not effective in rabies encephalitis. The use of tribavirin warrants further study, possibly combined with new therapeutic methods.