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PubMed2000

Efficient diagnosis of dengue infections using patients' peripheral blood leukocytes and serum/plasma.

OBJECTIVE: Dengue fever has been one of the most important health problems in Taiwan since a large outbreak during 1987 and 1988. It is critically necessary to have a diagnostic approach that can detect early infections in an outbreak or even find infections existing in silent transmission of the disease. METHODS: To develop an efficient diagnostic protocol, 105 plasma/serum and 35 peripheral blood leukocyte (PBL) specimens from the 1994 outbreak in southern Taiwan were collected for assessment by various diagnostic techniques in this study. RESULTS: In acute blood samples, dengue viruses were isolated from 19.4% (14/72) and 33.3% (14/42) of reported and confirmed cases, respectively. Viral RNA in serum/plasma was detected from 20.0% (12/60) of acute samples, which was significantly higher than that from convalescent samples (3/44; 6.8%). However, viral RNA in PBLs, detected by reverse transcription polymerase chain reaction (PBL-RT-PCR), could be observed in 73.2% (19/26) and 66.7% (6/9) of acute and convalescent samples, respectively. The persistence of dengue viruses in PBLs was also evidenced by the presence of viral antigens in 42.9% (4/7) of confirmed convalescent samples by the immunofluorescence antibody test. In addition, IgM antibodies were detected in 43.8% (46/105) of reported cases and 85.2% (46/54) of confirmed cases by the IgM antibody capture enzyme-linked immunosorbent assay (MAC-ELISA). CONCLUSIONS: Although IgM antibody detection achieved the highest detection rate among techniques assessed in this study, no individual test can actually reach full efficiency for early diagnosis of dengue infections. Here, we propose a protocol which applies MAC-ELISA and PBL-RT-PCR in sequence, by which 22 confirmed cases were definitely proved as dengue positive. High levels of both sensitivity and specificity were shown in this protocol.

Antibodies, Viral↗
PubMed1983

Characterization of circulating immune complexes in chronic non-A, non-B hepatitis.

We studied the frequency and composition of circulating immune complexes (CIC) in patients with chronic persistent non-A, non-B (NANB) hepatitis and in convalescent persons after an apparently normal recovery from acute NANB hepatitis. 10 of 16 patients with chronic NANB hepatitis and 5 of 11 convalescent persons after acute NANB hepatitis had CIC as detected by the Raji cell technique. CIC in chronic NANB hepatitis were composed of IgG, C3, and in 7 of 10 cases also IgM, while in CIC from convalescent persons IgG and C3 were present, too, but IgM was detected in only 1 case. Viral antigens within the CIC were not detectable in any case while 14 of 16 chronic NANB hepatitis patients were found to have free virus-associated antigen in serum.

Antigen-Antibody Complex↗
PubMed1997

High levels of circulating interleukin-4 and interleukin-10 in Kawasaki disease.

For analysis of the cytokine network in Kawasaki disease (KD), we measured over time the plasma levels of interferon (IFN)-gamma, interleukin (IL)-4 and IL-10 in patients with KD. Fifteen patients with KD were studied. Eight healthy children were selected as control subjects. Circulating IFN-gamma levels were measured by immunoradiometric assay, and IL-4 and IL-10 levels were measured by enzyme-linked immunosorbent assay. The results were as follows: (1) The plasma levels of IFN-gamma in KD patients in the acute phase were significantly higher than the levels of patients in the convalescent phase (p < 0.05) and those of the control children (p < 0.05). (2) The plasma levels of IL-4 in the KD patients in the acute phase were significantly higher than the levels of the patients in the convalescent phase (p = 0.001) and those of the control children (p = 0.001). (3) The plasma levels of IL-10 in the KD patients in the acute phase were significantly higher than the levels of the patients in the convalescent phase (p < 0.03) and those of the control children (p < 0.005). (4) The investigation of the relationship between the IL-4 and IFN-gamma levels during the acute phase of KD demonstrated a significant reciprocal relationship (p < 0.05). (5) There was no significant relationship between the IL-4 and IL-10 levels during the acute phase. However, plasma IL-10 levels were low in the patients with high levels of plasma IL-4, and the patients with high levels of IL-10 revealed low levels of plasma IL-4. The above results suggested that a variety of patterns of cytokine production was present in the acute phase of this disease, and that the key cytokine, which might regulate the cytokine network, was IL-4.

Case-Control Studies↗
PubMed1991

Bacterial infection and peripheral vascular disease.

Whole blood filterability was monitored in 16 nondiabetic peripheral vascular disease (PVD) patients within forty-eight hours of onset of bacterial infection, after ten to seventeen days antibiotic therapy and again, ten days later, after convalescence. The whole blood filterability rate was constantly disturbed before infection in these patients; the impairment worsened significantly (as was expected during infection), but after convalescence the whole blood filterability rate did not return to preinfection levels. This further significant impairment in whole blood filterability was inversely correlated with a reduction in the patients' pain-free walking time as determined by a standard treadmell test performed after convalescence and compared with their average times before infection.

Bacterial Infections↗
PubMed2004

Adrenomedullin is highly expressed in blood monocytes associated with acute Kawasaki disease: a microarray gene expression study.

Kawasaki disease (KD) is an acute inflammatory disorder of children frequently associated with the development of coronary artery abnormalities. Although a great deal is known about inflammatory and immune responses in acute KD, the mechanisms linking the immune response to vascular changes are not known. To gain further insight into this process, we performed a microarray gene expression analysis on RNA isolated from the peripheral blood mononuclear cells of four patients with KD during both their acute and convalescent phases. Forty-seven genes of 7129 genes examined showed an increased expression in three or all four patients in the acute compared with the convalescent phase of KD. Fourteen of these genes were significantly (p < 0.05) up-regulated, including several inflammatory response genes (e.g. S-100 A9 protein) and also anti-inflammatory genes (e.g. TSG-6). Of greatest interest, the adrenomedullin (ADM) gene, known to be associated with coronary artery vasodilation, was up-regulated in the acute phase of KD (p = 0.024). Up-regulation of ADM in the acute phase of KD was confirmed in peripheral blood mononuclear cells of 11 additional KD patients by reverse transcriptase-PCR (p < 0.01). Isolated blood monocytes but not lymphocytes were demonstrated by real-time PCR to have increased ADM mRNA (p = 0.01). Plasma ADM protein level in 32 additional KD patients was also confirmed to be higher in acute KD compared with convalescent KD (p < 0.032). It is interesting that from microarray results, other molecules known to be associated with coronary dilation, including nitric oxide, prostacyclin, acetylcholine, bradykinin, substance P, and serotonin, were not elevated in acute KD. Our current study suggests that ADM-expressing monocytes that infiltrate the coronary vascular wall may be the cause of coronary dilation in the acute phase of KD.

Acute Disease↗
PubMed1997

Diagnostic value of cerebrospinal fluid examination in children with peripheral facial palsy and suspected Lyme borreliosis.

Our objective was to determine the diagnostic value of CSF examinations in the diagnosis of neuroborreliosis in children with peripheral facial palsy (PFP). Paired serum and CSF samples from 21 children with PFP were investigated for antibody responses to Borrelia burgdorferi antigens using three different ELISA systems and one Western blot assay. Twenty of the children (95%) had detectable immunoglobin (Ig) M or IgG in the acute-phase serum, but discrepancies between serologic assays were noted in 33% for IgM and 22 to 50% for IgG. Intrathecal specific-antibody production was detected in five of the 20 seropositive children (25%). These five patients showed seroconversion in convalescent sera in at least one assay. Similar seroconversion suggesting recent infection with B. burgdorferi was observed in eight of the 10 children (80%) without intrathecal specific-antibody production, from whom convalescent serum samples could be obtained. All patients with intrathecal antibodies or seroconversion had shown lymphocytic pleocytosis in the acute phase of PFP. In the acute phase of PFP the detection of intrathecal production of antibodies to B. burgdorferi allows prompt diagnosis of neuroborreliosis. For patients with lymphocytic pleocytosis but no detectable intrathecal antibodies, analysis of convalescent serum may help to establish this diagnosis.

Adolescent↗
PubMed1986

Analysis of histopathologic findings in cases with dilated cardiomyopathy with special reference to formulating diagnostic criteria on the possibility of postmyocarditic change.

From our study employing serial endomyocardial biopsy in patients with acute viral or idiopathic myocarditis, we were able to construct histopathologic criteria for acute, subacute and convalescent myocarditis. We realize that it is difficult for the inexperienced observer to make an appropriate diagnosis of myocarditis or postmyocarditic changes in patients with dilated cardiomyopathy (DCM). In order to overcome this problem, each finding was graded and the scores obtained were analyzed statistically and compared with those from hypertrophic cardiomyopathy (HCM) and chronic right ventricular overloading (CRVO). The scores were obtained by summing the gradings for each of the following findings: increase of fibrocytes, increase of fibroblasts in the interstitium, hypertrophy of myocytes, fragmentation of muscle bundles, interstitial fibrosis, disarrangement of muscle bundles, abnormal branching, variation in size, increased glycogen deposition in the sarcoplasm, scarcity of myofibrils, and nuclear degeneration of myocytes and endocardial thickening. Since the increase in number of fibrocytes was considered important, its value was doubled. Scores in each group were as follows: convalescent myocarditis: 17.1 +/- 4.7 (n = 10), DCM: 13.2 +/- 3.3 (n = 47), HCM: 9.7 +/- 2.4 (n = 20), CRVO: 7.0 +/- 3.6 (n = 21). It was found that the scores for cases with myocarditis in the convalescent stage and in DCM were higher than those found for cases with either HCM and CRVO (p less than 0.05). In summary, the high score for postmyocarditis in DCM could suggest that prior myocarditis is an important causative factor of this disease.

Biopsy↗
PubMed2005

Complement levels in Brazilian children during and after meningococcal meningitis.

PURPOSE: To evaluate the functional activity of the classical and alternative pathways of the complement system and the levels of C3, C4, and factor B during the first episode of meningococcal infection and during the convalescence period. PATIENTS AND METHODS: Ten Brazilian children ranging in age from 8 months to 8 years, admitted from 1991 to 1993 with a clinical-laboratory diagnosis of meningococcal meningitis, were studied during acute infection (up to 7 days from diagnosis) and during the convalescence period (1 to 6 months after the acute episode). C3, C4, and Factor B were measured using nephelometry, and the lytic activity of classical and alternative pathways were evaluated by a kinetic method and expressed as the time needed to lyse 50% of an erythrocyte suspension (T1/2, expressed in seconds). Low T1/2 values for classical and alternative pathways correlate with high activities of the classical and alternative complement pathways, respectively. RESULTS: A significant difference was observed between the alternative pathway lytic activity during infection and the convalescence period (282 vs 238 seconds, respectively, P = .01). No differences were detected in the other complement parameters analyzed. CONCLUSIONS: In the presence of meningococcal meningitis, the alternative pathway is preferentially activated. This is probably due to the greater ability of the meningococcal endotoxin to activate this pathway in vivo.

Acute Disease↗
PubMedDate not supplied

Activated T cells and Leu-7+ cells in Bell's palsy.

Patients with Bell's palsy were investigated with regard to the cellular immune response during the acute and convalescent stages with the purpose of acquiring information concerning the etiology of the condition. In order to ascertain whether there was activation of the immune system, the proportion of activated T cells in the peripheral blood of 14 patients with Bell's palsy was analysed using monoclonal antibodies. A transient increase of these cells occurred in the acute stage, with a return to normal in the convalescent stage. The Leu-7+ cells which play a role in the natural cellular defence against viral infections, were also studied by using monoclonal antibodies in 25 patients. A significant change in the percentage of Leu-7+ cells was not found when the samples from the entire group were analysed. However, 10 patients who were investigated between September and October 1984, when the incidence of Bell's palsy was comparatively high, showed significantly lower percentages of Leu-7+ cells in the acute stage compared with the convalescent stage. The increase in the activated T cells in the acute phase of the palsy suggests a cell-mediated, immuno-regulatory abnormality with primary or secondary immune activation. Similar cellular immune alterations are found also in multiple sclerosis. Further, the differences in the proportions of Leu-7+ cells which occurred in these 10 patients may denote a contribution of various etiological factors to the disease.

Adult↗
PubMed1997

Evidence for selection of 11 amino acid CDR3 domains in V kappa III-derived immunoglobulin light chains in Kawasaki disease.

Kawasaki disease (KD) is a rheumatic disease that occurs during childhood. Although T cells have been implicated as having an important role in the pathogenesis of KD, the role of B cells is unclear. To detect preferential expression of 11 amino acid complementarity determining region (CDR)3 domains, we used two-stage PCR (polymerase chain reaction) to analyze the CDR3 lengths of VkIII-derived immunoglobulin kappa light chains expressed in peripheral blood B cells during the acute, subacute, and convalescent phase of this disease. As controls, the peripheral blood B cells of age-matched normal and children with acute febrile diseases other than KD were tested. In 5 of 7 KD patients, expression of kappa light chains containing 11 amino acid codon CDR3 intervals was increased during the acute and subacute phase of KD but decreased during the convalescent phase. Two of the 7 KD patients showed the same pattern during the subacute and convalescent phase, but not during the acute phase. Two of the 5 patients with acute febrile diseases other than KD showed increased expression of kappa chains with 11 amino acid codon CDR3 intervals, but it was not a major fraction. Three of the 5 patients with acute febrile diseases other than KD and all normal control subjects showed only 9 and 10 amino acid CDR3 domains. These results strongly suggest that B cells expressing kappa light chains with the 11 amino acid CDR3 domains might be involved in the pathogenesis of KD.

Acute Disease↗
PubMed2005

Gene expression profiles in peripheral blood mononuclear cells of SARS patients.

AIM: To investigate the role of inflammatory and anti-viral genes in the pathogenesis of SARS. METHODS: cDNA microarrays were used to screen the gene expression profiles of peripheral blood mononuclear cells (PBMCs) in two SARS patients (one in the acute severe phase and the other in the convalescent phase) and a healthy donor. In addition, real-time qualitative PCR was also performed to verify the reproducibility of the microarray results. The data were further analyzed. RESULTS: Many inflammatory and anti-viral genes were differentially expressed in SARS patients. Compared to the healthy control or the convalescent case, plenty of pro-inflammatory cytokines such as IL-1, TNF-alpha, IL-8, and MAPK signaling pathway were significantly upregulated in the acute severe case. However, anti-inflammatory agents such as IL-4 receptor, IL-13 receptor, IL-1Ra, and TNF-alpha-induced proteins 3 and 6 also increased dramatically in the acute severe case. On the contrary, a lot of IFN-stimulated genes like PKR, GBP-1 and 2, CXCL-10 and 11, and JAK/STAT signal pathway were downregulated in the acute severe case compared to the convalescent case. CONCLUSION: Gene expression in SARS patients mirrors a host state of inflammation and anti-viral immunity at the transcription level, and understanding of gene expression profiles may make contribution to further studies of the SARS pathogenesis.

Adult↗
PubMed1979

Absence of leukocytes permissive to dengue 2 virus in the acute phase of dengue hemorrhagic fever.

Patients with primary dengue infection developed dengue 2 virus (D2V) permissive peripheral blood leukocytes (PBL) 2--3 weeks after infection. PBL from healthy individuals with dengue antibody were permissive to D2V in vitro, suggesting that immunologically mediated in vitro D2V permissiveness persists for a relatively long time after recovery from dengue infection. However, PBL obtained from second infection dengue hemorrhagic fever patients did not support D2V growth during the acute phase of illness but did so during convalescence. Leukocytes from dengue-immune patients with typhoid fever or non-dengue viral illness were permissive throughout both acute and convalescent phases of illness although there was tendency for increased permissiveness during convalescence. Acute phase PBL from DHF patients synthesized and secreted dengue neutralizing antibody in culture. Absence of D2V replication in these cultures was strongly, but not completely, correlated with antibody production. Other immunological mechanisms, in addition to antibody, may be operating in vitro or in vivo during acute phase dengue hemorrhagic fever to alter the permissiveness of PBL to D2V infection.

Adolescent↗
PubMed1983

Specificities of antibodies boosted by acute Plasmodium falciparum infection in man.

In the search for antibodies correlating with host-protective immunity to Plasmodium falciparum in man, sera from individuals in Papua New Guinea were analyzed at the time of infection and in the convalescent period following infection. Titers of antibody were determined by enzyme linked immunoassay (ELISA), and the specificities of antibodies was examined by gel electrophoresis of immunoprecipitates. In the majority of cases, convalescence was associated with an increase in antibody titer and one-dimensional gel analysis of immunoprecipitated biosynthetically labeled parasite antigens demonstrated the variability in specificity of the antibody response in the two types of serum samples from different individuals. A protein of Mr 96,000 which has previously been identified as a candidate host-protective antigen was not clearly seen in immunoprecipitates generated with acute serum, even in samples with high titers of antibody assessed by ELISA. Antibodies to a protein Mr 96,000 were present in some, but not all convalescent sera. Two-dimensional gel analysis was more sensitive in detecting a boost in antibody response to minor antigens (e.g., an acidic protein Mr approximately equal to 230,000). This approach has not led to identification of antibody specificities to major antigens which are invariably boosted by infection and drug cure, but has identified antibody specificities in acute sera which are inadequate in quantity or quality to inhibit parasite growth.

Adult↗
PubMed1988

Lymphocyte responsiveness to a candidate malaria sporozoite vaccine (R32tet32) of individuals with naturally acquired Plasmodium falciparum malaria.

Lymphocyte proliferative responses to the candidate malaria sporozoite vaccine antigen R32tet32 were evaluated in 29 patients with acute Plasmodium falciparum malaria, 20 convalescent patients, 11 nonimmune individuals, and 22 healthy residents of two endemic malarious areas in Thailand. The results indicate that 14 of 20 (70%) convalescent patients and 14 of 22 (64%) residents of endemic areas responded to the R32tet32 antigen. However, only 8 of 29 (28%) patients with acute P. falciparum malaria responded. When 4 of the convalescent patients who remained in a malaria-free area were restudied 5-10 months after the acute infection, they were either not responsive or their responses had greatly diminished. These findings show that sensitization to R32tet32 occurs following a natural P. falciparum infection, but the cellular immune response to sporozoite antigens may be short-lived and may be suppressed during acute P. falciparum malaria.

Acute Disease↗
PubMed1991

Comparison of European isolates of viruses causing hemorrhagic fever with renal syndrome by a neutralization test.

Different virus isolates causing hemorrhagic fever with renal syndrome (HFRS) were compared using a neutralization test. Patient convalescent sera and antisera prepared in rabbits were used to compare Puumala-related Hantavirus isolates from Finland, Sweden, Belgium, and the USSR. The majority of European isolates were indistinguishable from each other using both homologous rabbit antisera and patient convalescent sera. The European isolates of HFRS were also compared with prototype Hantaan (the etiologic agent of Korean hemorrhagic fever). The one-way cross-reaction between the Hantaan and Puumala viruses, previously described using human convalescent sera tested by indirect immunofluorescence and immunoprecipitation, was also seen by the neutralization test.

Europe↗
PubMed2003

Transmission of epidemic dengue hemorrhagic fever in easternmost Indonesia.

In April 2001, a second suspected outbreak of dengue hemorrhagic fever in the easternmost region of Indonesia was investigated in Merauke, a town located in the southeastern corner of Papua, by the Indonesian Ministry of Health and the U.S. Naval Medical Research Unit No. 2. Principal case criteria of hemorrhagic disease provided for a study enrollment of 15 clinically acute and 37 convalescing subjects. Additionally, 32 comparable age/sex controls were selected from neighboring households. Laboratory diagnosis involved three testing methodologies: virus isolation by cell culture, a reverse transcriptase-polymerase chain reaction (RT-PCR) assay, and serologic assays. Antibody (IgM) to dengue virus was detected in 27% of the acute clinical cases, 30% of the convalescing cases, and only 3% of the matched controls. Dengue 3 was the only viral serotype detected from acute serum samples by the RT-PCR. The mean +/- SD age of the acute and convalescing cases was 7.8 +/- 5.4 years. Overall hospital records accounted for 172 suspected outbreak cases, all urban residents of Merauke with no recent travel history outside the area. The estimated outbreak-associated case fatality rate among all suspected dengue cases was 1.2%. A seven-year retrospective review of hospital records in Merauke showed negligible disease reporting involving hemorrhagic disease prior to the outbreak.

Adolescent↗
PubMed1979

A high molecular weight antigen in Legionnaires' disease bacterium: isolation and partial characterization.

We isolated a high molecular weight antigen of the Legionnaires' disease (LD) bacterium by column chromatography. The antigen was composed of 35% carbohydrate, 2.6% protein, 1.8% phospholipid, and 1% 2-keto-3-deoxyoctonate and was important in the host's antibody response because it inhibited the indirect immunofluorescent and microagglutination titers of convalescent sera from patients with Legionnaires' disease. The antigen also formed precipitin bands with seven of 10 convalescent sera from patients with Legionnaires' disease. We found chemical and biological evidence of endotoxinlike activity associated with the antigen. Cell sonicates and acid extracts of the LD bacterium gave multiple bands in immunodiffusion with human convalescent serum and rabbit antisera prepared against heat-killed LD bacteria. The antigenic structure of the LD bacterium therefore appears complex.

Animals↗
PubMed1981

Antibody response to structural proteins of measles virus in patients with natural measles and subacute sclerosing panencephalitis.

By immunoprecipitation and SDS-polyacrylamide gel electrophoresis, antibody responses to the structural proteins of measles virus were examined on patients with various forms of natural measles, atypical measles, and subacute sclerosing panencephalitis (SSPE). The serum of atypical measles most strongly reacted with four structural proteins, i.e., hemagglutinin (H), nucleocapsid (NC), fusion (F), and matrix (M) proteins. In natural measles, antibodies to the four structural proteins were detected at such an early convalescent stage as one month after the onset of disease. In late convalescent serums taken 9 years after natural measles, however, only low level antibody to M protein was present, whereas antibodies to H, NC and F proteins persisted. The serums and cerebrospinal fluid of SSPE patients showed patterns similar with those of the late convalescent serums. In Vero cells infected with cell-associated SSPE viruses (Niigata-1, ZH, and SI strains), M protein was not clearly demonstrated with serum of either atypical measles or SSPE patients, whereas H protein was demonstrated.

Adult↗