Enzyme immunoassays (EIAs) for the detection of anti-Haemophilus ducreyi serum IgA, IgG, and IgM antibodies.

BACKGROUND AND OBJECTIVES: Chancroid is a risk factor for heterosexually acquiring HIV. Controlling its spread may reduce HIV transmission. GOAL OF THE STUDY: To develop EIAs for assessing antibody levels and for seroepidemiologic studies. STUDY DESIGN: Anti-Haemophilus ducreyi IgA, IgG and IgM EIAs were standardized using a crude cocktail antigen. Evaluation was on sera from Kenya, Rwanda, Thailand and The Gambia. The two-tailed student's t test was used to compare results. RESULTS: The specificity of IgA was 97% (95% confidence interval (CI): 95-99%), of IgG was 92% (95% CI: 89-95%), and of IgM was 99% (95% CI: 98-100%). The sensitivity of IgA was 88% (95% CI: 83-93%), of IgG was 93% (95% CI:89-97%), and of IgM was 78% (95% CI:71-85%) in patients having an ulceration for more than eight days. Thus, 95% (95% CI:92-98%) of the chancroid patients were seropositive for at least one antibody type. The IgG and IgA EIAs were more sensitive in patients older than 24 years of age. Higher IgG rates were found in HIV infected chancroid patients. CONCLUSION: The EIAs should be useful for studying the kinetics of antibody levels and the epidemiology of H. ducreyi infection.

PIP: In Belgium, the Department of Infection and Immunity of the Institute of Tropical Medicine in Antwerp modified an experimental enzyme immunoassay (EIA) for the detection of serum IgG to Hemophilus ducreyi to develop EIAs for detection of anti-H. ducreyi IgA and IgM antibodies. They tested the modified EIA on sera from people in Nairobi, Kenya; Kigali, Rwanda; Banjul, The Gambia; and Bangkok, Thailand, who had a sexually transmitted disease. The EIA was able to identify correctly those who did not have anti-H ducreyi IgA, IgG, and IgM antibodies in 97%, 92%, and 99% of cases, respectively. Among people with a genital ulceration for more than 8 days, it was able to identify correctly those who had IgA, IgG, and IgM antibodies in 88%, 93%, and 78% of cases, respectively. 95% of all culture-proven chancroid patients tested seropositive for at least 1 antibody type. The sensitivity of IgG and IgA EIAs was significantly enhanced in patients with culture-proven chancroid who were older than 24 years old (p .01). HIV seropositive people from Kigali who had culture-proven chancroid had higher anti-H. ducreyi IgG seropositivity rates (but not IgA and IgM seropositivity rates), than did HIV seronegative chancroid people from Kigali (p .05). The increased IgG seropositivity rate was not related to higher antibody titers, however, suggesting that HIV infection modifies the response to H. ducreyi. These results show that the 3 EIAs hold promise as a means to study the kinetics of antibodies and the epidemiology of chancroid.

Eradication of hantavirus infection among laboratory rats by application of caesarian section and a foster mother technique.

Hantavirus antibodies were demonstrated by the indirect immunofluorescent antibody assay, in the serum of inbred strains of laboratory rats, during the period 1973-1982, at the Unit of Experimental Immunology in the Catholic University of Louvain, Brussels, Belgium. LOU rats, as well as immunocytomas, which were requested by laboratories in the U.K. and The Netherlands, were supplied at a time when the infection was unknown and unsuspected in Europe. Hantavirus-infected laboratory rats were rendered free of virus through re-derivation by caesarian section and suckling by virus-free foster mothers. Immunocytomas were tested for the presence of hantaviruses by implantation into seronegative laboratory rats. The strain of hantavirus causing the laboratory infection was clearly different from the one circulating in free-living bankvoles in Belgium. The exchange of laboratory rats and rat tumours in relation to the potential risk of laboratory-acquired hantavirus infection, is discussed.

HIV antigen detection in circulating immune complexes.

Various methods were evaluated for their effectiveness in releasing HIV antigen (Ag) from artificial immune complexes (IC) and from IC present in serum from HIV antibody (Ab) positive subjects. The most effective methods for recovering HIV Ag from IC were those which included a denaturation step to prevent reassociation of Ag with Ab. IC precipitation in 2.5% polyethylene glycol followed by acid treatment with 1 M glycine.HCl (pH 2) for 10 min at 70 degrees C in the presence of 0.05% SDS gave very satisfactory results. With this method, IC were detected in sera from HIV antibody positive Caucasian subjects at all stages of infection. After HIV IC dissociation, HIV Ag was detected in a significant number (8/17 or 47%) of asymptomatic subjects. IC were most prevalent during the late stages of infection. A substantial increase in HIV Ag positivity was also observed in 20 Senegalese HIV Ab positive sera. After HIV IC dissociation HIV antigen detection increased from 2/20 to 12/20. The relevance of IC detection is discussed.

Evidence for Hantavirus disease in Slovenia, Yugoslavia.

In Slovenia, North-Western part of Yugoslavia, 17 clinically documented Hantavirus disease cases (HVD) were serologically confirmed so far. Previously HVD was reported in the Southern part of Yugoslavia. By the indirect fluorescent antibody test (IFA), the prevalence of IgG class antibodies against different Hantaviral antigens was demonstrated in human sera collected in Slovenia. Three different reactivity patterns were observed. Majority of the IFA-positive human sera were confirmed by the immunoblot method. The distribution of Hantaviral infections was examined in small mammals captured in two natural foci of HVD, where clinical documented cases were reported. Hantaviral antibodies and antigens were demonstrated in C. glareolus, A. flavicollis, A. sylvaticus, and M. musculus.

Comparison of immunofluorescence and enzyme-linked immunosorbent assays for the serology of Hantaan virus infections.

Three enzyme-linked immunosorbent assay (ELISA) systems based upon different principles were developed for the serology of Hantaan virus infections and compared with an indirect immunofluorescence assay (IFA). The indirect IFA was carried out with gamma-irradiated Hantaan virus-infected and uninfected Vero E6 cells fixed with ethanol (-70 degrees C) or acetone (20 degrees C) on drop slides and a FITC-coupled sheep anti-human Ig preparation. Atypical staining in the IFA was avoided by using ethanol (-70 degrees C) instead of acetone (20 degrees C) fixation. In the first ELISA ('cell-assay'), Hantaan virus-infected or uninfected Vero E6 cells were used as antigens, which after gamma-irradiation were seeded into microtiter ELISA strips. Serial dilutions of human sera were incubated and specific antibodies were demonstrated with a horseradish peroxidase (HRPO)-conjugated sheep anti-human Ig preparation. In the second ELISA ('competition-assay') an affinity-purified human Ig preparation was used as a capture antibody for Hantaan virus antigen. After incubation of serial dilutions of human sera with this coat, the reactivity of the affinity purified anti-Hantaan virus Ig coupled to HRPO was determined. In the third ELISA ('complex trapping blocking [CTB]-assay') the same capture antibody was used to react with a mixture of the antigen and serial dilutions of human sera. The reactivity with the same HRPO conjugate was then determined. The results obtained in the respective assay systems with sera from people at risk or suspected of Hantaan virus infection coincided well. The CTB-ELISA proved to be faster and more sensitive than both the other ELISA systems, without giving more non-specific reactions: it detected almost all the IFA positive samples.(ABSTRACT TRUNCATED AT 250 WORDS)

Partial characterization of a Hantavirus isolated from a Clethrionomys glareolus captured in Belgium.

A Hantavirus was isolated in Vero-E6 cells from lungs of a free living bank vole (Clethrionomys glareolus) captured in Turnhout, Province of Antwerp--Northern part of Belgium. With help of monoclonal antibodies the Belgian Hantavirus isolate could be clearly differentiated from Hantaan virus strain 76-118, Prospect Hill virus strain PH1 and SR11, a Hantavirus isolated from laboratory Wistar rat in Japan, but not from the nephropathia epidemica virus strain Hällnäs.