Evaluation of a new Epstein-Barr virus Combi Test for rapid serologic diagnosis of infectious mononucleosis.

We evaluated the EBV Combi Test (Virion) in serum samples from 574 children with a clinical presentation suggestive of infectious mononucleosis (IM) and compared its performance with several other EBV serological tests. Out of 574 sera 66 gave an acute IM pattern, 406 gave a past infection pattern and 102 were found negative in the EBV Combi Test. Positive VCA IgM and VCA IgG IFA results, in the absence of EBNA antibodies, were found in 62 cases in which the EBV Combi Test gave an acute IM pattern. In addition, 4 to the 574 tested sera gave an acute positive result in the EBV Combi Test (two of them were VCA IgM positive and the other two VCA IgM negative but also EBNA negative). None of these four sera were CMV IgM or Toxoplasma gondii IgM positive. The heterophile antibody test was positive in only 28, and VCA IgM EIA positive in 44 of the 62 IM cases. These data confirm the necessity for an EBV serological diagnosis in children where the clinical diagnosis of EBV infectious mononucleosis must be confirmed or ruled our.

[The complement fixation reaction: a traditional, updated method for the detection of antibodies in the diagnosis of infections]. / Reakcija vezanja komplementa: tradicionalna, osuvremenjena metoda za otkrivanje protutijela u dijagnostici infekcija.

Complement-fixation (CF) is still an important basic serologic test for the diagnosis of infectious diseases. In several areas of microbiology (viral, bacterial, parasitic and fungal) it has served as a reference standard against which other methods have been compared. Its partial displacement by other techniques, as advocated in some recent literature, is often unfounded and uncritical; displacement is mostly due not to problems inherent in the method, but rather to the lack of reagents of satisfactory quality. The CF technique has been greatly improved in recent years. Improvements include the replacement of reaction tubes with microtiter 96-well plate systems, the availability of semi-automated and automated pipetting devices, the wide range of commercially offered antigens (over 60, by far larger than the range of antigens available for other test systems), an extreme stability of freeze-dried reagents with shelf-lives over decades and, last but not least, low reagent costs (particularly if compared with some other methods). For some diseases CF is still the method of choice (Campylobacter jejuni, Neisseria gonorrhoeae, Mycoplasma pneumoniae, Influenza A and B and some other respiratory viruses). CF has been recognized as a confirmatory test for Lyme disease (B. burgdorferi) and, probably, for Legionellosis. It has been used routinely for the follow-up of antibiotic treatment in syphilis and, recently, in Helicobacter pylori infections. A positive CF result is also indicative for the treatment of complications in Entamoeba histolytica carriers.(ABSTRACT TRUNCATED AT 250 WORDS)

[Asymptomatic varicella infections in pregnancy: detection and problems]. / Asymptomatische Varizelleninfektionen in der Schwangerschaft: Nachweis und Problematik.

Two cases of asymptomatic varicella infection during pregnancy will be presented. The diagnosis was made by the evidence of varicella-zoster-virus (VZV)-specific IgM or by the significant increase in antibody titer. The incidence of such subclinical VZV infections must be the issue of further investigations. Because of the rare occurrence of an embryopathy no general screening is usually made. On the other hand serological testing is recommended in cases of VZV infection in the surroundings of a pregnant woman. The problem of false-positive enzyme immunoassays make it necessary to confirm the results with one, or better two, additional methods.

Development of a recombinant enzyme-linked immunosorbent assay for detection of antibodies against Epstein-Barr virus nuclear antigens 2A and 2B.

The baculovirus expression system was used to produce full-length Epstein-Barr virus nuclear antigens (EBNAs) 2A and 2B. Recombinant baculoviruses that contained the EBNA-2A- and EBNA-2B-encoding sequences were constructed. The proteins were expressed in Spodoptera frugiperda SF-9 cells infected with the recombinant viruses and were characterized by using monoclonal and human polyclonal antibodies by immunoblotting and immunofluorescence techniques. Partially purified extracts of the EBNA-2A- and EBNA-2B-infected insect cells were used to establish a new enzyme-linked immunosorbent assay for the detection of antibodies against EBNA-2A and EBNA-2B. Preferential reactivity toward the type A or type B EBNA-2 protein was observed in 36% of serum specimens from Swiss patients with acute infectious mononucleosis and in 81% of Swiss patients with latent Epstein-Barr virus infection. Of the patients in the latter group, sera from 76% reacted preferentially with EBNA-2A, sera from 5% reacted preferentially with EBNA-2B, sera from 12% showed similar reactivities against both antigens, and sera from 7% were nonreactive.

A new Combi test for simultaneous detection of antibodies to viral capsid, early and EBNA antigens of Epstein-Barr virus.

In order to facilitate the differentiation between a recent (acute) and a past Epstein-Barr virus (EBV) infection, the Combi test was developed. This test is an anticomplement immunofluorescence test (ACIF) requiring only a single serum dilution to be tested on a single cellular spot. The cell line used expresses viral capsid antigen (VCA) and early antigen (EA) in about 5 to 10 percent of the cells as well as EBV nuclear antigens (EBNA) in more than 90 percent of cells. A satisfactory agreement between the Combi test and other tests for antibodies to EBV was obtained (IgG and IgM antibodies to VCA by IFA and EIA and antibodies to EBNA by ACIF including tests for heterophile and complement-fixing antibodies). When the standard serological tests gave negative results, the Combi test was also negative (absence of any fluorescence in the cells). Serologically confirmed recent (acute) infections lead to specific fluorescence in only 5 to 10 percent of the cells, while past infections result in fluorescence in 90 percent or more of the cells. For the diagnosis of a reactivated EBV infection or of EBV-associated malignancies, other tests should be employed. The test is based on the measurement of the activation and specific distribution of the C3 component of complement; the antibody class differentiation is therefore not necessary. The presence of rheumatoid factor (RF) and the IgG competition phenomenon do not influence the results of the Combi test. An introduction of the Combi test will enable a simplified, less expensive and more reliable serodiagnosis of EBV infections.

[The combined immunoenzyme test: anti-HIV-1 and -2 and anti-HTLV-1 in the detection of donors with retrovirus infections]. / Ispitivanje kombiniranog enzimskog imunoloskog testa: anti-HIV 1/2 i anti HTLV-1 u otkrivanju davatelja zarazenih retrovirusima.

Serums of whole blood donors, plasma donors, hemophiliacs, persons with risks behavior and normal population simultaneously were tested for markers of infectious diseases, anti-HIV-1 with Plivazim and anti-HIV-1/2 and anti-HTLV-1/2 with Roche Retrovirus EIA. The positive results were confirmed by immunofluorescence assay and Western blot. Nonspecific reactive serums were detected by Roche Retrovirus EIA and by Plivazim EIA, but there was no significant difference in the frequence of reactive results. Roche Retrovirus EIA test had specificity of 99.16% and sensibility of 97.56% as compared to Plivazim. Simultaneous testing of donors with a combined test for anti-HIV-1/2 and anti-HTLV-1 is equally reliable as testing with only anti-HIV-1.

[False rubella diagnosis in pregnancy by IgM enzyme immunoassay]. / Falsche Rubella-Diagnose in der Schwangerschaft aufgrund eines IgM-Enzymimmunoassays.

On the basis of a case report of a false-positive rubella diagnosis in early pregnancy the interpretation of positive IgM is discussed. A primigravida at 14 weeks of gestation showed rubella-specific IgM in an enzyme immunoassay. This result could not be confirmed by an alternative method (hemagglutination-inhibiting test after separating the serum by density gradients into different immunoglobulin classes and removal of the IgG with protein A). The immunological status corresponded with a past rubella infection or with the status after vaccination (IgG-positive). It is therefore recommended to use an additional method in the presence of IgM, especially if an interruption of the pregnancy has to be considered. In the course of the examination a recent cytomegalovirus infection could be detected.

[The clinical spectrum of rickettsiosis in the area of Zadar]. / Klinicki spektar rikecioze na zadarskom podrucju.

The clinical spectrum of rickettsioses in patients admitted to our hospital (Department of Infectious Diseases, General Hospital of Zadar) from 1983 to 1988 has been analysed. This spectrum consisted of the following diseases: murine typhus, Q-fever, boutonneuse fever and Brill-Zinsser disease. The low number of hospitalized patients with rickettsioses was in disproportion with the prevalence of antibodies to these rickettsiae in general population of the Zadar area. Our results indicate that the main reason for this disproportion are misdiagnosed and undiagnosed diseases.

Murine typhus on the northern Dalmatian islands, Yugoslavia.

The present epidemiological study had the objective to establish whether and to what extent resident islander populations might be affected by the natural foci of murine typhus. To this end, 294 serum specimens were collected during summer and autumn of 1985 from the northern Dalmatian islanders, all of which were tested for anti-Rickettsia typhi (R. typhi) complement fixing (CF) antibodies. These were detected and confirmed at both stages of the screening, namely during the first stage, conducted on the northern Dalmatian islands, where 63.3% of the representative population had a titre of 1:4 or higher and during the second stage, carried out on the island of Vir, with 68.1% of the residents found R. typhi positive unlike in the former. The majority of positive sera were found in the 21-60-year group, peaking in the 21-30- and 31-40-year groups with fewer positive sera among the subjects over 60 and the fewest among the residents below 20 years of life. Thus, current differences in antibody rates in various age groups were shown to be statistically significant. Murine typhus immunity in resident islander populations could be detected from the age of 10 years on, and was shown to become almost equal to the adult population's positive sera levels in the 16-20-year age group. No differences were revealed with regard to the infective agent exposure between the male and female populations representative of the total population or in any of the age groups.