Maroto Mc

Detection of mycobacterial DNA in papulonecrotic tuberculid lesions by polymerase chain reaction.

Tuberculids are a heterogenous group of cutaneous lesions. Recent discoveries of M. Tuberculosis DNA in these lesions by PCR suggest that M. tuberculosis could play a role in their pathogenesis. The aim of this study was to demonstrate the presence of M. tuberculosis DNA by polymerase chain reaction in papulonecrotic tuberculid lesions. Skin biopsy specimens from ten patients with papulonecrotic tuberculid lesions (histopathologic features) were studied. All of them tested solidly positive in a tuberculin intradermal test. A gene‐amplification PCR, using primers capable of amplifying DNA in the M. tuberculosis complex, was performed to detect M. tuberculosis DNA in the lesions. A 285‐bp sequence specific of M. tuberculosis complex was amplified and confirmed by Southern‐blot hybridation with a 32 p 5′‐labelled internal probe. No inhibitors were detected in the negative PCR samples. The PCR technique makes the detection of mycobacterial DNA in tuberculids a possibility, and therefore provides a rational basis for antituberculous therapy and for the clinical management of these disorders. J. Clin. Lab. Anal. 14:133–135, 2000.

ELISA test to detect Chlamydophila pneumoniae IgG.

A new ELISA test (Chlamydophila pneumoniae IgG, Vircell, Spain) to detect Chlamydophila pneuoniae IgG was evaluated. The micro‐immunofluorescence (MIF) test was used as reference method. Chlamydia trachomatis and Chlamydophila psittaci elementary bodies were also assayed. Two hundred and sixteen sera were included in the study: 66 from patients with peripheral arterial occlusive disease (Panel 1), 68 from adults with pneumonia (Panel 2), 44 from healthy adults (Panel 3) and 38 from patients with a sexuality transmitted disease by C. trachomatis (Panel 4). In Panel 1, 51 sera (77%) had antibody titres between 32 and 128; 4 out of 15 sera with IgG titres <32 were positive by ELISA test and 2 sera with 32 IgG titres were uncertain by ELISA; the remaining 60 sera were correctly classified, giving a 91% concordance between the techniques. In Panel 2, 55 sera (81%) had IgG titres between 32 and 512; 2 out of 13 sera with IgG titres <32 were positive by ELISA and 2 sera with 32 titres were uncertain by ELISA; the remaining 64 sera were correctly classified, giving a 97% concordance. In Panel 3, 22 sera (50%) had IgG titres between 32 and 64; only 1 out of 22 sera with IgG titres <32 was positive by ELISA, giving a 97% concordance between the techniques. In Panel 4, there were 24 (63%) negative, 10 (26%) uncertain and 4 (10%) positive results by ELISA, giving an 86% concordance. The C. pneumoniae ELISA test demonstrated 100% sensitivity and 85% specificity. The IgG ELISA test demonstrated a good concordance with the MIF test without the drawbacks associated with the latter assay. We conclude that the ELISA test could be an alternative to the MIF test.

Reliability of low-avidity IgG and of IgA in the diagnosis of primary infection by rubella virus with adaptation of a commercial test.

The detection of IgA and low‐avidity IgG and antibodies in serum is a potentially useful marker of recent infection by a microorganism. We studied the reliability of IgG avidity and presence of IgA for the diagnosis of recent acute infection by rubella virus. Low‐avidity IgG (Avy‐EIA test) was determined with a modified commercial test using 8 molar urea (indirect ELISA, DiaSorin, Italy) and IgA was determined with a homemade indirect ELISA test. Twenty‐five patients with recent primary infection by rubella virus (group I) and 50 healthy subjects (group II) were studied. In group I low‐avidity IgG varied between 100 and 0% (67.3 ± 21.8%); IgA was present in 24 patients (96%). In group II low‐avidity IgG varied from 50.4 to 0% (19.8 ± 16.9%). IgA was present in 2 subjects (4%). The sensitivity of the Avi‐EIA and the IgA test was 92 and 96%, respectively; specificity was 100 and 96%, respectively. We conclude that both low‐avidity IgG and IgA tests are helpful and reliable for the diagnosis of recent primary infection. J. Clin. Lab. Anal. 13:1–4, 1999.

Seroepidemiological study of hepatitis E virus in different population groups

In order to determine the seroprevalence of hepatitis E virus, 1,993 sera (453 from healthy pregnant women, 491 from Moroccan subjects, 492 from blood donors, 321 from children, and 236 from intravenous drug users) were studied. IgG was measured by enzyme immunoassay (EIA), and positive results were confirmed by Western blot. The EIA detected antibodies in 3.96 % of the subjects (5.6 % of the Moroccans and drug users and 1.8 % of the children). Fifty-four percent of these results were confirmed by Western blot, 11.4 % were found to be negative, and 34.2 % indeterminate. The overall prevalence after confirmation by Western blot decreased to 2.15 %. When studying the Western blot pattern of the positive samples, 95 % showed antibodies to SG-3, 65 % to 8–5, and only 9.3 % to CKS fusion protein. In the indeterminate Western blots, the results for these proteins were 96.3 %, 62.9 %, and 37 %, respectively. When the epidemiological data were analysed, no statistically significant differences between women and men or between different age groups were found.

Prevalence and viral persistence of TT virus in patients on hemodialysis.

Abstract The aim of this study was toevaluate the prevalence of TT virus (TTV) in 107 hemodialysis patients and 100 healthy individuals. Sixteen percent of hemodialysis patients and 2% of the control population were TTV positive (P<0.001). Sex, time on dialysis, transfusions, and alanine aminotransferase levels were not related to the presence of TTV DNA. TTV was more frequent in patients 50 to 70 (12/51) years of age and persisted in the serum of six of nine patients 8 months later. TTV was found in peripheral blood mononuclear cells of all patients with chronic infection, and sequence variation was noted in the peripheral blood mononuclear cells of two patients. No relationship between TTV infection and hepatitis was observed. Viral persistence and nonparenteral transmission may be possible in patients on hemodialysis.

Relationship between viral genotype and viral load in patients with chronic hepatitis C

To determine the distribution of hepatitis C virus (HCV) genotypes and to relate genotype to viral load, genotyping and quantification of viral RNA were carried out in 35 patients with chronic hepatitis C. Subtype 1a was most prevalent (43%), followed by subtypes 1b (23%) and 3a (14%). Mean viral load (log HCV-RNA copies/ml) for subtypes 1b, 1a and 3a was 7.1 ± 1, 5.6 ± 1.1 and 4.1 ± 2.4, respectively. The presence of immunoglobulin M was related to the duration of hepatitis and genotype 1 to a more severe hepatic injury and a higher viral load. Differences observed in viral load for a single HCV subtype justify the need to quantify HCV-RNA prior to establishing antiviral therapy.

Human immunodeficiency virus type-1 can be detected in monocytes by polymerase chain reaction.

Lymphocytes and monocytes from 25 patients infected with human immunodeficiency virus type-1 (HIV-1)--13 asymptomatic, seven with the AIDS-related complex (ARC) and five with the acquired immunodeficiency syndrome (AIDS)--were lysed and subjected to PCR with three primer pairs: SK38/SK39 (gag), SK68/SK69 (env) and SK29/SK30 (LTR). Amplified DNA was solution-hybridised with 32P-labelled probes (SK19, SK70 and SK31, respectively) and detected by PAGE-autoradiography. HIV-1 DNA was detected as follows. Asymptomatic patients: monocytes--gag 61.5%, env 100%, LTR 0%; lymphocytes--gag 100%, env 92.3%, LTR 53.84%. ARC patients: monocytes--gag 71.4%, env 57.1%, LTR 0%; lymphocytes--gag 100%, env 71.4%, LTR 71.4%. AIDS patients: monocytes--gag 80.0%, env 100%, LTR 0%; lymphocytes--gag 100%, env 60%, LTR 60%. The presence of HIV-1 DNA was confirmed in the monocyte fraction. In this cell subset, the env gene-directed primers were the most effective for amplification, whereas the LTR gene-directed primers failed to amplify HIV-1 DNA. The different pattern of amplification found in monocytes may suggest that these cells could be infected by a genetic variant of the virus.

Immune complex p24 antigen: A new prognostic marker in human immunodeficiency virus infection

p24 antigen (p24 Ag) is usually not found in patients in the asymptomatic phase of HIV infection, mainly because it is bound to anti-p24 antibodies (p24 Ab), with which it forms immune complexes. Nine asymptomatic patients positive for HIV-1 antibodies (stage II CDC criteria) treated with rα-interferon were followed (7.5+/-0.5 months), and a basal and final sample were tested for p24 Ag immune complexes (p24 Ag-IC), free p24 Ag, p24 Ab, CD4 count, β2-microglobulin, properdin B factor, and C3 and C4 complement fractions. p24 Ag-IC was detected in 55.5% of the samples, whereas free p24 Ag was detected in 5.5%. The finding of no change or an increase in p24 Ag-IC serum levels may be considered a prognostic marker. We believe that p24 Ab cannot be used as a prognostic marker unless p24 Ag-IC detection is simultaneously evaluated. In der asymptomatischen Phase der HIV-Infektion läßt sich das p24 Antigen (p24 Ag) meist nicht nachweisen; dies ist vor allem darauf zurückzuführen, daß es an anti-p24 Antikörper (p24 Ab) gebunden ist und mit ihnen Immunkomplexe bildet. Bei neun anti-HIV-1 positiven, asymptomatischen Patienten (Stadium II nach CDC Kriterien), die mit r-alpha-Interferon behandelt wurden, wurden Verlaufsbeobachtungen (7,5 +/- 0,5 Monate) durchgeführt. Basis- und Abschlußuntersuchungen auf p24 Ag Immunkomplexe (p24 Ag-IC), freies p24 Ag, p24 Ab, CD4 Zellzahlen, β2-Mikroglobulin, Properdin B Faktor und Komplementfraktionen C3 und C4 wurden dabei vorgenommen. In 55,5% der Proben fand sich p24 Ag-IC, in 5,5% freies p24 Ag. Die Beobachtung gleichbleibender oder ansteigender Werte von p24 Ag-IC im Serum hat möglicherweise prognostische Bedeutung. Unserer Ansicht nach kann der p24 Ab nicht als prognostischer Marker eingesetzt werden, wenn nicht zugleich die Bestimmung von p24 Ag-IC erfolgt.Summaryp24 antigen (p24 Ag) is usually not found in patients in the asymptomatic phase of HIV infection, mainly because it is bound to anti-p24 antibodies (p24 Ab), with which it forms immune complexes. Nine asymptomatic patients positive for HIV-1 antibodies (stage II CDC criteria) treated with rα-interferon were followed (7.5+/-0.5 months), and a basal and final sample were tested for p24 Ag immune complexes (p24 Ag-IC), free p24 Ag, p24 Ab, CD4 count, β2-microglobulin, properdin B factor, and C3 and C4 complement fractions. p24 Ag-IC was detected in 55.5% of the samples, whereas free p24 Ag was detected in 5.5%. The finding of no change or an increase in p24 Ag-IC serum levels may be considered a prognostic marker. We believe that p24 Ab cannot be used as a prognostic marker unless p24 Ag-IC detection is simultaneously evaluated.ZusammenfassungIn der asymptomatischen Phase der HIV-Infektion läßt sich das p24 Antigen (p24 Ag) meist nicht nachweisen; dies ist vor allem darauf zurückzuführen, daß es an anti-p24 Antikörper (p24 Ab) gebunden ist und mit ihnen Immunkomplexe bildet. Bei neun anti-HIV-1 positiven, asymptomatischen Patienten (Stadium II nach CDC Kriterien), die mit r-alpha-Interferon behandelt wurden, wurden Verlaufsbeobachtungen (7,5 +/- 0,5 Monate) durchgeführt. Basis- und Abschlußuntersuchungen auf p24 Ag Immunkomplexe (p24 Ag-IC), freies p24 Ag, p24 Ab, CD4 Zellzahlen, β2-Mikroglobulin, Properdin B Faktor und Komplementfraktionen C3 und C4 wurden dabei vorgenommen. In 55,5% der Proben fand sich p24 Ag-IC, in 5,5% freies p24 Ag. Die Beobachtung gleichbleibender oder ansteigender Werte von p24 Ag-IC im Serum hat möglicherweise prognostische Bedeutung. Unserer Ansicht nach kann der p24 Ab nicht als prognostischer Marker eingesetzt werden, wenn nicht zugleich die Bestimmung von p24 Ag-IC erfolgt.

Clinical utility of a competitive ELISA to detect antibodies against Treponema pallidum.

Screening for Treponema pallidum infection is carried out on a large human population. To reduce costs, fewer tests which still offer adequate sensitivity and specificity could be performed. We studied the reliability of a novel indirect ELISA method to test for this infection. Several panels of sera were used that corresponded to 40 primary infections (group 1), 13 recurrences (group 2), 348 latent infections (group 3), 5 samples with anticardiolipin antibodies (group 4), 15 samples from patients with Lyme borreliosis (group 5), and 400 samples from blood donors and healthy pregnant women (group 6). The ELISA showed a global sensitivity and specificity of 100 and 99.5%, respectively. Our evaluation shows that Enzygnost Syphilis is a sensitive, specific, and simple test to screen for this infection. J. Clin. Lab. Anal. 14:83–86, 2000.

Gbv-C RNA presence in several high-risk groups of Spain

GB virus C subtype (GBV-C) seems to share the same routes of transmission as other parenteral transmitted viruses. We have evaluated the prevalence of GBV-C in 247 patients with potential risk for GBV-C infection and in 91 healthy blood donors. The presence of GBV-C RNA was examined by polymerase chain reaction in serum samples. The 23.6% of parenteral drug users were GBV-C positive, 36.3% of them were also HIV infected. Moreover, 22.5 and 19% of sera from patients with HBV and HCV chronic hepatitis, respectively, but without apparent risk factors seemed GBV-C infected. Finally, the 6% of patients on hemodialysis were also positive. Therefore, these results suggest that GBV-C is transmitted by parenteral routes but other non-parenteral routes shared with HBV or HCV must be considered.