Serosurveillance for double infection with Pseudomonas pseudomallei in tuberculous patients.

The serosensitivity to Pseudomonas pseudomallei antigen in pulmonary tuberculous patients was surveyed in Ubon Ratchathani Province, a melioidosis-endemic and tuberculosis-prevalent area in Thailand. Indirect hemagglutination assay (IHA) and indirect immunofluorescent assay (IFA) for IgM and IgG were employed for the measurement of antibody levels, with cut-off points set at 1:160, 1:8, and 1:32, respectively. Retrospective protocol survey of clinical data was also conducted. From these studies, however, no evidence was obtained to show that tuberculous patients have a disposition to acquire double-infected with P. pseudomallei and to develop melioidosis. The serosensitivity was never higher than that of the general population of the province as represented by healthy blood donors, nor related with the clinical severeness. Tuberculosis and melioidosis appear to be mutually independent diseases without showing interrelated prevalence in the endemic area.     

Serological diagnostics of imported malaria in Czechoslovakia

Two serologic techniques for malaria detection were compared in this study; the indirect fluorescent antibody (IFA) test used in 214 persons (38 Czechoslovak citizens returning from visits to tropical countries and 176 foreign visitors arriving to Czechoslovakia from areas endemic for malaria) and the indirect hemagglutination (IHA) test employed in 125 persons (29 Czechoslovak citizens and 96 foreigners). Comparisons revealed poor correlation between the IFA test and IHA test data. Of the two tests the IFA test appeared to be distinctly more reliable, more sensitive and more specific, the IHA test turned out to yield both false positive and false negative results. The antigen from Plasmodium gallinaceum gave lower IFA titres than P. falciparum antigen, but reacted with antibodies to all species of human plasmodia, and gave reliable test results. Positive serologic responses were appreciably more frequent in foreigners (46.0%) than Czechoslovak citizens (23.7%). The maximum percent positivity for malarial antibody was among individuals from tropical countries of Africa (74.6%), seropositivity in people from malaria endemic areas in Asia and Latin America was far less frequent (28.4% and 44.4%, respectively).     

Comparative serology techniques for the diagnosis of Trypanosoma cruzi infection in a rural population from the state of Querétaro, Mexico

Immunological diagnostic methods for Trypanosoma cruzi depend specifically on the presence of antibodies and parasitological methods lack sensitivity during the chronic and “indeterminate” stages of the disease. This study performed a serological survey of 1,033 subjects from 52 rural communities in 12 of the 18 municipalities in the state of Querétaro, Mexico. We detected anti-T. cruzi antibodies using the following tests: indirect haemagglutination assay (IHA), indirect immunofluorescence assay (IFA), ELISA and recombinant ELISA (rELISA). We also performed Western blot (WB) analysis using iron superoxide dismutase (FeSOD), a detoxifying enzyme excreted by the parasite, as the antigen. Positive test results were distributed as follows: ELISA 8%, rELISA 6.2%, IFA and IHA 5.4% in both cases and FeSOD 8%. A comparative study of the five tests was undertaken. Sensitivity levels, specificity, positive and negative predictive values, concordance percentage and kappa index were considered. Living with animals, trips to other communities, gender, age, type of housing and symptomatology at the time of the survey were statistically analysed using SPSS software v.11.5. Detection of the FeSOD enzyme that was secreted by the parasite and used as an antigenic fraction in WBs showed a 100% correlation with traditional ELISA tests.     

Schistosomiasis: Evaluation of the indirect fluorescent antibody,complement fixation,and slide flocculation tests in screening for schistosomiasis

Foreign Service personnel undergo pertinent parasitologic examinations upon return from foreign duty posts. Under this program, 2800 sera have been evaluated for schistosomiasis in this laboratory. The majority of individuals tested were considered to have limited exposure to schistosomiasis, although a few indigenous people from endemic areas also were screened. Nonindigenous populations usually gave stronger serological reactions than did indigenous populations. A comparison was made between those having protozoan and helminthic infections and those that were negative parasitologically. A number of subjects with tissue-phase helminths were evaluated and consistently gave strong reactions in the indirect fluorescent antibody (IFA) tests. On the other hand, there was no characteristic pattern observed in individuals with low serum titers. The IFA test proved to be highly sensitive and sufficiently specific for screening, provided that low background reactions were disregarded (i.e., when +/? and 1+ reactions were ignored at low serum dilutions). Thus, the IFA test was the method of choice for screening. Recourse to the complement fixation (CF) and slide flocculation (SF) tests, however, was necessary for definitive diagnosis. In view of the differences in the antigens and the serodiagnostic technics used in this survey, absolute correlation of test results could not be expected. Nevertheless, the three procedures (IFA, CF, and SF) showed excellent correlation in proven cases of schistosomiasis.     

Screening for Chagas disease by immunoenzymology: comparison of ELISA with immunofluorescence and indirect hemagglutination in 976 blood donors

The efficiency of an immunoenzymatic technique (ELISA) for the systematic research of Chagas' disease in blood donors was compared with one of 2 well-known methods, indirect haemagglutination (IHA) and indirect fluoro immuno assay (IFA). For the ELISA technique two different antigenic extracts from epimastigote culture forms of T. cruzi, were used for sensitizing the polystyrene plates: a crude extract (Ag R) and a delipidized one (Ag B). Firstly the authors tested these 3 techniques in 5 control groups: sera from Chagas' disease, negative control sera, sera from visceral leishmaniasis, african trypanosomiasis and finally monoclonal gammapathies, the high levels of blood proteins being a possible cause of false positives. Secondly the screening of Chagas' disease was performed in the same way in 976 blood donors from Recife, Brazil. In the case of the Ag-R extract used in the ELISA technique a high cross-reactivity was found with visceral leishmaniasis sera, along a risk of false positives with gammapathic ones. The sensitivity of this technique was found to be high (3,3 +/- 1 p. cent of positive blood donors) and a very good correlation was found with the reference techniques, IFA and IHA, the sensitivity of which is lower (2,3 +/- 1 and 1,7 +/- p. cent). The use of a delipidized antigenic extract (Ag B) for the ELISA technique is not suitable, in spite of an apparent higher specificity: indeed, the positives rate is high (11,5 +/- 0,2 p. cent), but the correlation is very weak or non existent in the case of IHA or IFA. In conclusion, the ELISA technique using a crude extract of T. cruzi appears to be a very convenient method for screening blood donors with Chagas' disease, the lack of specificity due to leishmaniasis or monoclonal blood proteins not posing any real problem to blood banking.     

A preliminary survey for human immunodeficient virus (HIV) infections in tuberculosis and melioidosis patients in Ubon Ratchathani, Thailand.

A preliminary survey was conducted for the prevalence of HIV infections in pulmonary tuberculosis and melioidosis patients in Ubon Ratchathani province, in Thailand, the second largest province in population which supplies labors to Bangkok metropolis. In this province, tuberculosis is prevalent in a higher rate than in most other provinces and melioidosis is endemic. Four HIV-seropositives were found in a total of 551 suspected and culture-positive cases of pulmonary tuberculosis, while no HIV-seropositive was found in 121 melioidosis patients. In view of the rapidly expanding HIV-infections in Thailand, a strict watch will be needed on the future epidemiological status of HIV-infection in tuberculous patient.     

Serodiagnosis of cilia-associated respiratory bacillus infection by the indirect immunofluorescence assay technique

Antibody to cilia-associated respiratory (CAR) bacillus was detected by the indirect immunofluorescence assay (IFA) technique using tracheal sections of infected mice as antigen in serum samples collected from rats infected naturally and experimentally. Nine of 23 cases of natural infection were positive in IFA antibody, with titres ranging from 1:10 to 1:80, and all these antibody-positive cases were also histologically positive. The remaining 14 cases were negative in both IFA antibody and histological diagnosis, even though some of them were infected with Sendai virus and Mycoplasma pulmonis. In the experimental infection, serum samples collected from 18 rats on days 4, 7, 14, 21, 28 and 56 post-inoculation (PI) (three rats for each point) and examined for IFA antibody revealed that seroconversion occurred in one rat on day 14 PI and in three rats on day 21 PI. Antibody titres of 1:80 to 1:160 remained to the termination of the experiment. The IFA technique was useful for the diagnosis of CAR bacillus infection except in the early stage of the infection.     

Humoral antibody response and Ig characterization of the specific agglutinins in rabbits during experimental American trypanosomiasis

A comparative follow up study of the specific agglutinins detected by direct agglutination (DA) test and the immune response detected by specific lysis (SL), indirect immunofluorescence (IFA), indirect hemagglutination (IHA) and complement fixation (CF) tests in rabbits inoculated with trypomastigotes of T. cruzi is reported here.The specific antibody response was detected first by DA test. Reductive cleavage of sera with 2-mercaptoethanol produced a drop in the agglutinin titer of the sera during the first 30 days of infection.The next test to become positive was SL and later on the IFA, IHA and CF tests became positive simultaneously.When fractions obtained by column chromatography in Sephadex G-200 were tested serologically it was demonstrated that specific antibodies were detected mainly in fraction I (IgM) of the pooled rabbit sera obtained 15 days after inoculation (acute stage), and in fraction II (IgG) of the pooled sera obtained from rabbits 90 days after inoculation (chronic stage).Antigens prepared with trypsinized and formolized epimastigotes of three T. cruzi strains, belonging to each one of the different immunological groups described, worked similarly in the detection of specific agglutinin antibodies.Trypanosoma cruzi agglutinins were highly specific in their reaction with their homologous T. cruzi antigens as was proved by the low agglutinin titer obtained in sera from infected rabbits when, instead of T. cruzi epimastigotes, promastigotes of L. donovani were used as antigen, and by the incapacity of this parasite to absorb the T. cruzi agglutinins.     

Evaluation of enzyme linked immunosorbent--assay for the detection of anticysticercus antibodies in cerebrospinal fluid from patients with neurocysticercosis.

Enzyme linked Immunosorbent Assay (ELISA) was done for the detection of antibodies to Cysticercus cellulosae in 135 cerebrospinal fluid (CSF) and 152 serum samples from patients suspected clinically of neurocysticercosis (NC), neurological disorders other than NC and controls by the use of crude cyst extract antigen. This assay was compared with the standard technique of indirect haemagglutination test (IHA). The results of the two techniques were matched with retrospective analysis of proven diagnosis of these patients. ELISA and IHA was found to be positive respectively in 88 and 84 percent of CSF and 92 and 87.2 percent of serum samples from proven NC patients. The IHA technique was found to be absolutely specific for the detection of antibodies in CSF samples while cross reactions were observed with ELISA technique in CSF from 5 patients, one each suffering from disappearing CT scan lesion, tubercular meningitis (culture negative), chronic meningitis, benign intracranial hypertension and non compressive myelopathy. However possibility of neurocysticercosis cannot be absolutely ruled out in such patients. Both the techniques were found to be highly non specific for the detection of antibodies in serum samples. The study suggests that either of the two techniques may be used for the detection of antibodies in CSF samples from clinically suspected NC patients with high degree of sensitivity and specificity.     

Cat scratch disease. Survey on the presence of Bartonella henselae among cats of Tuscany

To verify the presence of Bartonella henselae-infection in cats living in Tuscany (central Italy) serological and bacteriological surveys were carried out. The blood serum samples of 427 cats, 254 living in private houses and gardens and 173 in public or private catteries, were tested for anti-B. henselae antibodies by indirect immunofluorescence assay (IFA). Among these samples, 35 were examined by IFA to detect antibodies against Bartonella quintana. Bacteriological examinations were performed on the blood samples, collected in EDTA (ethylene diaminetetraacetic acid), of 18 cats (10 seropositive to B. henselae and 8 negative). From each of the same 18 specimens DNA was extracted and used as template in polymerase chain reaction (PCR). The primers p24E and p12B were employed in the PCR assay to amplify a 296 bp fragment of the Bartonella 16S rRNA gene. IFA detected 98 (22.95%) B. henselae-positive serum samples (40-40.82% from cats living in houses and gardens and 58-59.18% from cats of catteries) at different antibody titers (70 at 1:64 titer, 4 at 1:128, 22 at 1:256, 2 at 1:512). Among the 35 sera tested to detect antibodies against B. quintana, 9 (25.71%) resulted positive at 1:64 titer; all these samples showed higher antibody titers to B. henselae. Out of the 26 negative sera, 20 were negative to B. henselae too and 6 had antibodies against B. henselae at 1:64. Hemocultures gave negative results. PCR scored positive with DNA of 4 B. henselae-seropositive cats, two of which belonged to two children with cat scratch disease (CSD).     

Review of the oriental leafhopper genus Lampridius (Hemiptera: Cicadellidae: Deltocephalinae), with description of a related new genus

The leafhopper genus Lampridius Distant, 1918 (type species: L. spectabilis Distant, 1918 ) is redescribed, and a second species, L. cuspidatus sp. nov. (Thailand: Loei), is described. A related new genus Paralampridius gen. nov. and four new species are described: P. mimicus sp. nov. (China: Guangdong, Hainan), P. rotundatus sp. nov. (Thailand: Loei), P. sinuatus sp. nov. (Thailand: Loei, Suphanburi, Petchaburi), and P. truncatus sp. nov. (Thailand: Chaiyaphum, Ubon Ratchathani). Both genera are tentatively included in the tribe Opsiini, although only Lampridius has paired aedeagal shafts with separate gonopores. All six species are illustrated, and a key is provided for their identification. Problems with the current tribal classification are discussed in light of the present discovery of closely related species that show variation in characters previously assumed to be stable at the tribal level in the subfamily Deltocephalinae.     

The serodiagnosis of human hydatid disease: 2. Additional studies on selected sera using indirect haemagglutination (IHA), enzyme linked immunosorbent assay (ELISA) and defined antigen substrate spheres (DASS).

Sixty-one serum samples selected on the basis of reactivity in the complement fixation (CF) and latex agglutination (LA) test, were further examined for sensitivity and specificity by indirect haemagglutination (IHA), enzyme linked immunosorbent assay (ELISA) and defined antigen substrate spheres (DASS). Twenty sera from healthy Europeans and 48 samples from patients with either schistosomiasis or trichinosis were also tested. Comparable levels of sensitivity were found between the CF and LA positive sera and IHA, ELISA and DASS. Of the CF positive LA negative group of sera, many were positive by DASS but only a few reacted in IHA and ELISA. Some cross reactivity was also observed in the schistosomiasis sera tested by IHA and ELISA.     

Serological Diagnosis of Human Melioidosis with Indirect Hemagglutination and Complement Fixation Tests

An indirect hemagglutination (IHA) test and a complement fixation (CF) test were evaluated from test results on sera from 212 human melioidosis patients of which 119 were culturally proved cases. Significant antibody titers (IHA titers of 1:40 or greater and CF titers of 1:4 or greater) were demonstrated with either test in all except five patients. IHA and CF titers ranged as high as 1:20,480 and 1:1,024, respectively. Antibodies were usually demonstrated by both tests 1 week after onset of disease. Transient seronegative reactions during the course of disease were seen in sera of approximately 19% of the patients with either IHA and CF but rarely with both tests. High titers in either test were obtained by the third week of disease and reached maximum levels in 4 to 5 months. Titers usually were detectable for 9 or more months. Antibodies were detected by IHA and CF tests in 80 to 100% of the sera obtained at various time intervals from 9 months to 2 or more years after disease onset. Antibody persistence occurred in patients who had a short disease course, as well as in patients with prolonged, complicated infections. The IHA test had excellent specificity when evaluated with normal human sera and diverse antimicrobial sera from hyperimmunized rabbits and human patients. The CF antigen appeared to contain common antigens with some but not all types of Pseudomonas aeruginosa. The specificity of the CF antigen could be enhanced without appreciable effect on its sensitivity by use of a titer of 1:8 in lieu of 1:4 as a criterion for a significant reaction. Either test could be used advantageously for the laboratory diagnosis of melioidosis.     

An indirect ELISA of classical swine fever virus based on quadruple antigenic epitope peptide expressed in <Emphasis Type="Italic">E.coli</Emphasis>

In this study, a synthesized quadruple antigenic epitope gene region of the classical swine fever virus (CSFV) E2 glycoprotein was expressed in E. coli to a obtain target protein. This target protein was used as a coating antigen to establish an indirect ELISA for specifically detecting anti-CSFV antibodies in serum samples from pigs. The P/N cut-off value of this assay was 1.92 by receiver operating characteristic curve (ROC) analysis based on 30 negative sera and 80 positive samples. The test gave 97.5% sensitivity and 96.7% specificity compared with the indirect hemagglutination (IHA) test. The inter-assay and intra-assay coefficients of variation (CVs) for 16 sera were both ⩽6.8%. No cross-reactivity between the coating antigen and anti-bovine viral diarrhoea virus (BVDV) antibodies was observed.     

Evaluation of an indirect hemagglutination inhibition technique for identifying antibody in animals experimentally and naturally exposed to fungi

Rabbits were exposed to aerosols containing spores of Aspergillus fumigatus, Aspergillus flavus, Aspergillus niger, or of a Penicillium sp. Sera from these rabbits were tested by indirect hemagglutination (IHA) and by IHA inhibition. The serologic reactions with the rabbit sera were compared to reactions with sera from cattle naturally exposed to airborne microorganisms. By three months of age, most cattle had positive IHA reactions to A. fumigatus and Penicillium antigens. The IHA inhibition tests indicated that antibody production in 12 of the 20 cattle probably resulted from exposure to A. flavus. One calf reacted as if sensitized by A. niger. Two were totally nonreactive. Five of the cattle had reactions that were not identifiable relative to the reactions in rabbits.Purchased by U.S. Department of Agriculture for Official Use.     

Serological survey for two simian retroviruses in macaques and African green monkeys

Colonies of nonhuman primates at the Bowman Gray School of Medicine (BGSM) were tested for antibodies to two retroviruses associated with immunodeficiency by indirect immunofluorescence (IFA) and western blot. A total of 471 cynomolgus macaques (Macaca fascicularis), 144 rhesus monkeys (M. mulatta) and 67 stumptail monkey M. arctoides) were tested for SRV-1, and 152 African green monkeys (Cercopithecus aethiops) were tested for SIV. Of the macaques tested, 170 (36%) cynomolgus, 5 (3%) rhesus and 8 (12%) stumptails were positive for SRV-1 antibodies by IFA. Of the African green monkeys, 54 (36%) were IFA positive for SIV antibodies. A total of 143 African green monkeys tested by IFA also were tested by western blot. In the African green monkeys, the IFA had a positive predictive value of 98% and a negative predictive value of 96%. Of 176 IFA positive macaque sera tested by western blot, 49 (28%) were positive, 55 (31%) were considered equivocal (only one band, usually to p27 core protein), and 72 (41%) were negative.     

Anti-VP1 and anti-VP2 antibodies detected by immunofluorescence assays in patients with acute human parvovirus B19 infection

Acute human parvovirus B19 infection is followed by an antibody response to the structural proteins of the viral capsid (VP1 and VP2). We used 80 sera collected from 58 erythema infectiosum and 6 transient aplastic crisis patients to test IgM and IgG antibodies against these two proteins in an immunofluorescence assay (IFA) using Sf9 cells infected with recombinant baculovirus expressing either VP1 or VP2 antigen. Although less sensitive than IgM capture enzyme immunoassay using native antigen (MACEIA), we could detect anti-VP1 or anti-VP2 IgM antibodies by IFA in 49 patients with acute infection (76.6%). Detection of IgG anti-VP1 and anti-VP2 by IFA, however, was as sensitive as IgG detection by indirect enzyme immunoassay. By applying IgG avidity IFA to sera of the 15 IgM IFA negative patients we were able to confirm acute infection in further 12 cases by IFA. Overall, acute infection was confirmed by IFA in 61 (95.3%) of the 64 patients.     

Seroprevalence of Toxoplasma gondii infection in dogs in Sichuan Province, southwestern China

There is a lack of information concerning the prevalence of Toxoplasma gondii infection in dogs from southwestern China. In the present study, serum samples from 314 household dogs were collected from Wenchuan, Heishui, and Jiuzhaigou in Sichuan Province, southwestern China, in May and June 201; sera were assayed for T. gondii antibodies using an indirect haemagglutination test (IHA). Antibodies to T. gondii were found in 11 of 314 (3.5%), with IHA titers of 1:64 in 4 dogs, 1:128 in 3, 1:256 in 2, 1:512 in 1, and 1:1024 in 1. No regional difference was observed among the 3 counties (P > 0.05). The results of the present study indicated that infection with T. gondii in dogs is common in China, including household dogs in Sichuan Province, and should be of public health concern.     

Detection of antibodies to cytomegalovirus, herpes simplex virus and rubellavirus with the micro-enzyme immunoassay.

Serum samples of pregnant women (200), non-pregnant women (100) and children (36) were tested for the presence of antibodies to cytomegalovirus (CMV), herpes simplex virus (HSV) and rubellavirus with the micro-enzyme immunoassay (micro-EAI), the indirect immunofluorescence antibody (IFA) technique and the haemagglutination-inhibition (HI) test. Micro-EIA gave the highest positivity rate: 78% for CMV, 86% for HSV and 85% for rubellavirus compared to 67% (CMV) and 84% (HSV) of the IFA technique and 79% (rubellavirus) of the HI test, respectively. Among IFA and HI positive sera the occurrence of micro-EIA negative ones was 6% for CMV, 4% for HSV and 9% for rubellavirus. It is concluded that the introduction of micro-EIA will improve the diagnostic and screening work of virus laboratories.     

Serotyping of Avian Mycoplasma Species in India

Two-hundred and two strains of avian mycoplasma species belonging to 10 biotypes were typed serologically by employing disk growth inhibition (DGI) and indirect hemagglutination (IHA) tests. These could be placed into seven serotypes, namely A (80), B (50), C (3), E (34), L (13), P (4), and 1 and R (18). The figures in parentheses show the number of strains within each type. A close relationship was observed between DGI and IHA tests. The IHA test, however, was more sensitive and specific. It was also noticed that biochemically identical biotypes, namely E and G, and B and M were also found identical in serotyping, thus confirming the biochemical identity. In view of these facts, the strains of biotypes M and G were grouped under serotypes B and E, respectively. The antigenic relationships between the serotypes are also discussed.