Toxoplasma gondii antibodies in HIV and non-HIV infected Thai pregnant women

Serological evidence for Toxoplasma gondii infection in Thai pregnant women was investigated. One thousand six hundred and sixty-nine blood specimens were collected from 838 HIV-seropositive and 831 HIV-seronegative pregnant women attending the antenatal-care clinic at Siriraj Hospital, Bangkok, Thailand, during a two-year period. Toxoplasma IgG antibody was detected, using a solid-phase enzyme-linked immunosorbent assay in which the membrane protein p-30 was the predominant antigen. IgG positive sera were subsequently examined for IgM antibody by the capture antibody enzyme immunoassay. The IgG antibody was found in 450 (53.7%) HIV seropositive women and 44 (5.3%) non-HIV infected women, with a statistically significant difference (p < 0.0001). Three of the 450 HIV-seropositive and 2 of the 44 HIV-seronegative sera with IgG antibody were positive for IgM antibody against T. gondii. This result suggested that HIV seropositive pregnant women had a higher risk of Toxoplasma infection with increase exposure to their offspring.     

Genotypic, immunophenotypic and clinical features of Thai patients with paroxysmal nocturnal haemoglobinuria

Paroxysmal nocturnal haemoglobinuria (PNH) is an acquired haematological disorder characterized by complement-mediated haemolytic anaemia caused by deficiency of glycosylphosphatidylinositol (GPI) anchored proteins. Somatic mutation of an X-linked gene, PIG-A, is responsible for the defect in biosynthesis of GPI-anchor. It appears that frequency of PNH differs geographically, and seems to be more frequent in some Asian countries, such as Thailand and China. We studied a group of 34 Thai patients with PNH to see whether the somatic mutations in PIG-A, extent of deficiency of GPI-anchored proteins (complete or partial) and complication with aplastic anaemia among Thai patients are different from those in other regions. We determined 37 PIG-A mutations in 33 patients (10 base substitutions, 14 single-base deletions, five multiple-base deletions, three single-base insertions, two multiple base insertions and three others) which were found to be similar to those found in European, American and Japanese patients. Most patients had cells with a complete deficiency of CD59 (type III cells), whereas 19% and 33% of the patients with reliable data for CD59 expression had partially deficient granulocytes and erythrocytes (type II cells), respectively. Most mutations resulted in a complete loss of function of PIG-A in accordance with the prevalent PNH III phenotype. 19 patients (51%) had aplastic anaemia; their PIG-A mutations were not different from those without pre-existing aplastic anaemia. These characteristics of Thai patients are similar to patients from other regions. There was some negative correlation between mean basal Hb concentration and percentage of CD59-negative granulocytes (r = -0. 374; P = 0.0476). In addition, patients with severe anaemia (basal Hb <7 g/dl) had a significantly higher percentage of affected granulocytes than those with mild anaemia (88.5 +/- 9.4 v 64.9 +/- 25.9; P = 0.01). The data suggest that the severity of anaemia in PNH depends partly on the size of the PNH clone.     

Immunophenotypic discrepancies between granulocytic and erythroid lineages in peripheral blood of patients with paroxysmal nocturnal haemoglobinuria

Abstract: In paroxysmal nocturnal haemoglobinuria (PNH), somatic mutation of the PIG‐A gene is thought to result in altered expression of glycosylphosphatidylinositol (GPI)‐anchored proteins. This study was performed to determine if there were any heterogeneities of cellular phenotypes between two major peripheral blood cells, erythrocytes and granulocytes. Using CD59‐based immunocytometry, the patterns of CD59 expression were shown to be conserved in the circulating erythroid cells (reticulocytes and mature erythrocytes) in all 29 patients with PNH. Twenty‐one patients had distinct combinations of PNH type I, II, and III cells in different lineages. Only eight patients exhibited similar patterns of CD59 expression between the two lineages. Approximately one third of the patients had PNH type II cells in either or both of the two lineages indicating variable lineage involvement. The proportion of abnormal granulocutes was higher than those of abnormal reticulocytes and erythrocytes. In patients with appropriate erythropoietic responses to haemolysis (RPI>2.0), shift reticulocytes display predominantly PNH phenotypes. These immature erythroid cells with altered expression of GPI‐anchored proteins may dominate the peripheral blood during periods of increased marrow activity resulting in greater phenotypic mosaicism in such patients. Discrepancies in expression of GPI‐anchored proteins in PNH which are highly variable between the two lineages may be the result of their different life spans and the influence of complement‐mediated cytolysis. The phenomena also indicated the possible occurrence of more than one PNH clones with variable clonal dominance.     

Intestinal parasitic infections among human immunodeficiency virus-infected and -uninfected children hospitalized with diarrhea in Bangkok,Thailand

A prospective observational study was conducted to determine the prevalence and the clinical impact of intestinal parasitic infections in diarrheal illness among HIV-infected and HIV-uninfected children hospitalized with diarrhea in Bangkok, Thailand. Stool samples were examined for intestinal parasites using a simple smear method, a formalin-ether concentration method, a modified acid-fast stain and a modified trichrome stain. Intestinal parasites (IP) were identified in the stool specimens of 27 of 82 (33%) HIV-infected and 12 of 80 (15%) HIV-uninfected children (p=0.01). Microsporidia and Cryptosporidium were the most common IP found. Eighty-two percent of HIV-infected and 97% of HIV-uninfected groups presented with acute diarrhea and 76% of each group had watery diarrhea. Pneumonia was the most common concurrent illness, found in 22%. Clinical findings were unable to differentiate children infected with IP. Sixty-three percent of HIV-infected and 83% of HIV-uninfected children who had IP made a satisfactory recovery without specific anti-parasitic therapy. However, 9 children (7 HIV-infected and 2 HIV-uninfected) with persistent diarrhea who also had cryptosporidiosis and/or microsporidiosis did not respond to azithromycin and/or albendazole respectively. HIV-infected children with cryptosporidiosis were older and had more advanced HIV infection than those with microsporidiosis. Routine stool examination for IP should be considered due to the absence of clinical markers. The lack of effective therapy for the major IP found underscores the importance of preventive measures.     

Infections in E-beta thalassemia

Infection is a major complication and the leading cause of death in thalassemia, especially E-beta thalassemia. The spectrum of infections in E-beta thalassemia include mild and severe infections, therapy-related infections such as Yersinia enterocolitica infection associated with desferrioxamine (DFO) therapy, and transfusion-transmitted disease, as well as unique infections such as with pythiosis. Prospective studies in Thailand indicate that patients with E-beta thalassemia had more frequent episodes of both mild and severe infections. The former included upper respiratory tract infection, acute gastroenteritis, cutaneous abscess, and gingivitis. Severe infections occurred more commonly in patients with splenectomy and included septicemia, pneumonia, biliary tract infection, salmonellosis, and urinary tract infection. Responsible organisms were Escherichia coli (26%), Klebsiella pneumoniae (23%), Salmonella (15%), and Streptococcus pneumoniae (13%). Other organisms included Pseudomonas, Staphylococci, Burkholderia pseudomallei (melioidosis), and Aeromonas. Patients undergoing DFO therapy are at risk for Y. enterocolitica infection which may be localized to mesenteric nodes and tonsils or occur as a generalized form such as septicemia. Recently, we have seen a unique infection so-called vascular pythiosis. Patients usually presented with clinical features of vascular occlusion of lower limbs from ascending arteritis and thrombosis. The causative organism, Pythium insidiosum, is fungus-like, in the kingdom Stramenopila, and in the class Oomycetes. The mortality rate is high and the only effective treatment has been early amputation or possibly immunotherapy. The predisposing factors of infections in thalassemia include splenectomy, iron overload, anemia, and granulocyte dysfunctions. General management of infections in thalassemia consist of prevention, i.e., immunization with pneumococcal and hepatitis vaccines, oral penicillins especially in patients with splenectomy, removal of predisposing factors such as gallstones, iron overload, and appropriate antibiotics.     

Induction of zoospore formation in Thai isolates of Pythium insidiosum

Three human isolates of Pythium insidiosum De Cock 1987, the aetiologic agent of pythiosis, were induced for zoosporogenesis. The isolates were grown on Sabouraud glucose agar for 2 days before being inoculated into boiled spikelets of Axonopus compressus (Gramineae) and then into induction medium (IM). Optimum zoospore formation was demonstrated under the light microscope after incubation for 24 h at 37 degrees C in IM.     

Development of flow cytometry for detection and quantitation of red cell bound immunoglobulin G in autoimmune hemolytic anemia with negative direct Coombs test

About 2-10% of patients with warm-antibody autoimmune hemolytic anemia (WAIHA) exhibit a negative direct Coombs test (DAT), requiring more sensitive tests, including detection of RBC-bound immunoglobulins by flow cytometry, for diagnosis. In this study, the optimal conditions for detection and quantitation of RBC-bound IgG by flow cytometry were studied using blood samples from six patients with AIHA and two healthy individuals. Quantitation of RBC-bound IgG was performed using quantum simply cellular (QSC) beads coated with goat anti-mouse IgG antibodies. For detection of RBC bound IgG, a 60-minute incubation of all blood samples with 40 microl of 1:10 dilution of FITC-conjugated mouse anti-human IgG gave mean fluorescent intensity (MFI) values comparable to experiments using larger amounts or higher concentrations of the anti-human IgG. The acquired antibody binding capacity (ABC) values (or IgG molecules) for each QSC bead level, at 40 microl of 1:5 and 1:10 dilution of anti-human IgG for 60 minutes were close to the manufacturer-assigned ABC values. The IgG molecules per RBC in all six patients with positive DAT of 4+, 3+, 2+, 1+, trace and negative DAT were 31,725, 3,823, 1,753, 524, 260 and 88 respectively and in two healthy individuals with negative DAT they were 104 and 78.     

Using ‘collective omics data’ for biomedical research training

Systems‐scale molecular profiling data accumulating in public repositories may constitute a useful resource for immunologists. It is for instance likely that information relevant to their chosen line of research be found among the more than 90,000 data series available in the NCBI Gene Expression Omnibus. Such ‘collective omics data’ may also be employed as source material for training purposes. This is the case when training curricula aim at the development of bioinformatics skills necessary for the analysis, interpretation or visualization of data generated on global scales. But ‘collective omics data’ may also be reused for training purposes to foster the development of the skills and ‘mental habits’ underpinning traditional reductionist science approaches. This review describes a small‐scale initiative involving investigators, for the most part immunologists, having engaged in a range of training activities relying on ‘collective omics data’.     

Cross over placebo control trial of dilazep in beta-thalassemia/hemoglobin E patients

An attempt was made to find better symptomatic treatment for beta-thalassemia/hemoglobin E (beta-thal/Hb E) patients in order to reduce their blood demand. Oral administration of dilazep was prescribed for these patients and a clinical trial was conducted over a 2-year period as a cross over placebo control study. Seventeen beta-thal/Hb E patients were enrolled in the study. All of them received dilazep and placebo for 10 months at different periods of time and were taken care of by the same doctor throughout the study. The blood demand of the same patients during the period of receiving dilazep with the period of receiving placebo, was 1.5 +/- 1.8 U/10 months versus 2.2 +/- 2.6 U/10 months, respectively. Thus dilazep showed a benefit in decreasing the blood demand by about 50% although the results did not reach statistical significance (p = 0.1). There was a statistical difference in hemoglobin concentration of the patients receiving dilazep compared with placebo (p = 0.038). While receiving dilazep the mean +/- SD hemoglobin level was 5.82 +/- 0.8 g/dl, significantly higher than while receiving placebo (5.66 +/- 0.9 g/dl) (p = 0.038). The liver, and renal function tests, and cardiac enzyme levels of the patients showed no significant changes throughout the study. However, one case had a problem with bleeding following tooth extraction whilst receiving dilazep and needed 1 unit of blood transfusion. In conclusion, administration of dilazep to patients with beta-thal/Hb E increased the patients' hemoglobin and reduced their blood demand with few side effects.     

Safety and immunogenicity of an HIV subtype B and E prime-boost vaccine combination in HIV-negative Thai adults

ALVAC-HIV (vCP1521) and AIDSVAX B/E were evaluated in a phase 1/2 trial of human immunodeficiency virus (HIV)-negative Thai adults. Of 133 volunteers enrolled, 122 completed the trial. There were no serious vaccine-related adverse events, nor were there intercurrent HIV infections. Lymphoproliferative responses to glycoprotein 120 E were induced in 63% of the volunteers, and HIV-specific CD8 cytotoxic T lymphocyte responses were induced in 24%. Antibody responses increased in frequency and magnitude in association with the dose level of AIDSVAX B/E. Binding and neutralizing antibodies to the MN strain were induced in 100% and 98%, respectively, of the volunteers receiving 600 microg of AIDSVAX B/E, and such antibodies to E strains were induced in 96% and 71%, respectively, of these volunteers. This vaccine combination was well tolerated and was immunogenic, meeting milestones for advancement to phase 3 evaluation.