Evaluation of immunoglobulin G subclass antibodies against recombinant Fasciola gigantica cathepsin L1 in an enzyme-linked immunosorbent assay for serodiagnosis of human fasciolosis

A cystatin capture enzyme-linked immunosorbent assay (ELISA) using recombinant Fasciola gigantica cathepsin L1 antigen was developed to detect specific immunoglobulin G (IgG) subclass antibodies (IgG1, IgG2, IgG3, and IgG4) and was evaluated for its diagnostic potential for human fasciolosis. In an analysis of the sera of 13 patients infected with F. gigantica, 209 patients with other parasitic infections, 32 cholangiocarcinoma patients, and 42 healthy controls, the IgG4-ELISA gave the highest diagnostic values. The sensitivity, specificity, accuracy, and positive and negative predictive values of this method based on the detection of IgG4 antibody were 100%, 99.3%, 99.3%, 86.7%, and 100%, respectively. The results revealed that restricting the ELISA to the detection of specific IgG4 antibody enhanced the specificity and accuracy for the serodiagnosis of human fasciolosis.     

Dengue myocarditis

We report a 13-year-old boy who developed bradycardia and hypotension a day after recovery from dengue hemorrhagic fever. His electrocardiogram, during the bradycardia, showed a junctional rhythm with a rate of 50 beats/minute. This is the first reported case of sinus node dysfunction following dengue infection.     

Evaluation of human IgG subclass antibodies in the serodiagnosis of angiostrongyliasis

Immunoglobulin G subclass antibody (IgG1, IgG2, IgG3, and IgG4) responses to the rat lungworm, Angiostrongylus cantonensis, were analyzed using the immunoblotting technique in an attempt to further improve the sensitivity and specificity for the serodiagnosis of human angiostrongyliasis. Serum samples from patients with proven angiostrongyliasis and from clinically suspected cases of angiostrongyliasis with eosinophilic meningitis were tested. Sera from patients with other parasitic illnesses and from healthy volunteers were also analyzed. The results indicate that the immunoblotting used to detect IgG4 antibodies to the antigenic band of an approximate molecular mass of 29 kDa from young adult somatic extract of A. cantonensis is the most reliable test. It gives accuracy, sensitivity, specificity, and positive and negative predictive values of 89.2%, 75%, 95%, 85.7% and 90.4%, respectively. More importantly, the test can discriminate between human angiostrongyliasis, gnathostomiasis and cysticercosis, three diseases that produce eosinophilic meningitis.     

Comparison between specified percentage and fifth percentile criteria for spirometry interpretation in Thai patients

OBJECTIVES: The present study was conducted to determine the degree of agreement between the interpretation of spirometry using a specified percentage of predicted value (SPC) and the fifth percentile (FPC) as the cut off between normal and abnormal. METHODOLOGY: Spirometric values were collected for 1754 subjects attending the respiratory service at Siriraj Hospital between February 2000 and April 2001. These values included forced vital capacity (FVC), forced expiratory volume in 1 s (FEV1), FEV1/FVC, maximal mid-expiratory flow (FEF25-75%) and peak expiratory flow (PEF). A comparison of results between SPC and FPC was performed. The SPC cut-off values for FVC, FEV1, FEV1/FVC, FEF25-75% and PEF were 80% predicted, 80% predicted, 70%, 65% predicted and 80% predicted, respectively. The FPC cut-off values were calculated from reference equations for the Thai population. Inter-rater agreement was calculated as the kappa score. RESULTS: High kappa scores were obtained for FVC (0.84), FEV1 (0.88) and FEF25-75% (0.80). However, poor agreement was found for FEV1/FVC (0.72) and PEF (0.61). When the cut-off values for SPC were modified to 90% of predicted values for FEV1/FVC and to 65% of predicted values for PEF, agreement was substantially improved to 0.92 and 0.89, respectively. CONCLUSIONS: Interpretation by SPC using cut-off values of 80% predicted for FVC and FEV1 and 65% predicted for FEF25-75% resulted in good agreement with FPC. However, the SPC cut-off values for FEV1/FVC and PEF should be modified to 90% predicted and 65% predicted, respectively.     

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.     

Rapid detection of Opisthorchis viverrini and Strongyloides stercoralis in human fecal samples using a duplex real-time PCR and melting curve analysis

Human opisthorchiasis caused by the liver fluke Opisthorchis viverrini is an endemic disease in Southeast Asian countries including the Lao People’s Democratic Republic, Cambodia, Vietnam, and Thailand. Infection with the soil-transmitted roundworm Strongyloides stercoralis is an important problem worldwide. In some areas, both parasitic infections are reported as co-infections. A duplex real-time fluorescence resonance energy transfer (FRET) PCR merged with melting curve analysis was developed for the rapid detection of O. viverrini and S. stercoralis in human fecal samples. Duplex real-time FRET PCR is based on fluorescence melting curve analysis of a hybrid of amplicons generated from two genera of DNA elements: the 162 bp pOV-A6 DNA sequence specific to O. viverrini and the 244 bp 18S rRNA sequence specific to S. stercoralis, and two pairs of specific fluorophore-labeled probes. Both O. viverrini and S. stercoralis can be differentially detected in infected human fecal samples by this process through their different fluorescence channels and melting temperatures. Detection limit of the method was as little as two O. viverrini eggs and four S. stercoralis larvae in 100 mg of fecal sample. The assay could distinguish the DNA of both parasites from the DNA of negative fecal samples and fecal samples with other parasite materials, as well as from the DNA of human leukocytes and other control parasites. The technique showed 100% sensitivity and specificity. The introduced duplex real-time FRET PCR can reduce labor time and reagent costs and is not prone to carry over contamination. The method is important for simultaneous detection especially in areas where both parasites overlap incidence and is useful as the screening tool in the returning travelers and immigrants to industrialized countries where number of samples in the diagnostic units will become increasing.     

Molecular identification of Ascaris lumbricoides and Ascaris suum recovered from humans and pigs in Thailand,Lao PDR,and Myanmar

Parasitology Research - Ascaris lumbricoides is the largest roundworm known from the human intestine while Ascaris suum is an internal parasite of pigs. Ascariasis, caused by Ascaris lumbricoides,...     

Evaluating the potential impact of the new Global Plan to Stop TB: Thailand, 2004-2005

OBJECTIVE: WHO's new Global Plan to Stop TB 2006-2015 advises countries with a high burden of tuberculosis (TB) to expand case-finding in the private sector as well as services for patients with HIV and multidrug-resistant TB (MDR-TB). The objective of this study was to evaluate these strategies in Thailand using data from the Thailand TB Active Surveillance Network, a demonstration project begun in 2004. METHODS: In October 2004, we began contacting public and private health-care facilities monthly to record data about people diagnosed with TB, assist with patient care, provide HIV counselling and testing, and obtain sputum samples for culture and susceptibility testing. The catchment area included 3.6 million people in four provinces. We compared results from October 2004-September 2005 (referred to as 2005) to baseline data from October 2002-September 2003 (referred to as 2003). FINDINGS: In 2005, we ascertained 5841 TB cases (164/100 000), including 2320 new smear-positive cases (65/100 000). Compared with routine passive surveillance in 2003, active surveillance increased reporting of all TB cases by 19% and of new smear-positive cases by 13%. Private facilities diagnosed 634 (11%) of all TB cases. In 2005, 1392 (24%) cases were known to be HIV positive. The proportion of cases with an unknown HIV status decreased from 66% (3226/4904) in 2003 to 23% (1329/5841) in 2005 (P< 0.01). Of 4656 pulmonary cases, mycobacterial culture was performed in 3024 (65%) and MDR-TB diagnosed in 60 (1%). CONCLUSION: In Thailand, piloting the new WHO strategy increased case-finding and collaboration with the private sector, and improved HIV services for TB patients and the diagnosis of MDR-TB. Further analysis of treatment outcomes and costs is needed to assess this programme's impact and cost effectiveness.     

Neurotoxicity of Coscinium fenestratum stem, a medicinal plant used in traditional medicine.

Coscinium fenestratum is a common medicinal plant widely used in the Indochina region, but scientific data on its safety is very limited. This study aimed to observe the effect of this plant on neurotoxicity and neurobehavior. Oral administration of plant alcoholic extract at dosages of 5, 10 and 20 mg/kgBW for 14 days increased the rats body weight and decreased the neuron density in the cerebral cortex, hippocampus and striatum. The plant extract significantly increased stereotyped behavior in licking but did not cause anxiolytic activity, anti-depression, sensory motor co-ordination impairment and ataxia. It is concluded that the plant possesses neurotoxicity and is able to induce neurobehavioral changes in rats. Therefore, the application of this plant as either drug or supplementary food should be reconsidered.     

Increased serum levels of macrophage colony-stimulating factor (M–CSF) in α-and β-thalassaemia syndromes: correlation with anaemia and monocyte activation

Abstract: Serum levels of M–CSF were determined by an ELISA method in 29 and 34 patients with HbH disease (α₁/α₂ or α₁/HbCS) or β⁰-thal/HbE, respectively, in 28 haematologically normal subjects and in five patients with anaemia due to iron deficiency or myelodysplasia. In HbH disease and β⁰-thal/HbE, M–CSF concentrations were significantly higher than those in the normal subjects [986 ± 138 and 1385 ± 133, respectively, vs. 500 ± 33 pg/ml (mean ± SEM); p <0.01, and p <0.001, respectively]. By contrast, in patients with anaemia due to iron deficiency, M–CSF levels were within the normal range. In HbH disease and in β⁰-thal/HbE, M–CSF levels correlated inversely with mean basal Hb values (r = –0.39, p = 0.05 and r = –0.60, p <0.001, respectively). In addition, in some of the HbH and β⁰-thal/HbE patients, monocyte ADCC activities towards red cells were tested and found to be approximately twice as high as those in normal controls [38.3±5.7 and 30.7 ± 4.6 vs. 17.8 ± 1.8 % specific lysis (mean ± SEM), respectively; p <0.01 and p <0.02, respectively]. When thalassaemic patients and normal controls were considered together there was a significant correlation between M–CSF levels and monocyte ADCC activities (r = 0.51, p <0.02). The results suggest that in HbH disease and in β⁰-thal/HbE, raised serum M–CSF contributes to the anaemia by enhancing the effector function of mononuclear phagocytes towards red cells.