Jukka Suni

CMV infection is usually associated with concurrent HHV-6 and HHV-7 antigenemia in liver transplant patients

Human herpesvirus 6 and 7 (HHV-6, HHV-7) have been recently reported in liver transplant patients. HHV-6 may cause fever, neurological disorders and hepatitis. The clinical significance of HHV-7 is less clear. HHV-6 and -7 are closely related to cytomegalovirus (CMV), and interactions between the viruses have also been suggested. In this study, we investigated the post transplant HHV-6 and -7 antigenemia was in relation to symptomatic CMV disease after liver transplantation. Consecutive 34 adult liver allograft recipients transplanted during 1999-2000 were included in the study. CMV infections were diagnosed by the frequent monitoring of pp65-antigenemia and by viral cultures. HHV-6 and -7 were demonstrated, by using immunoperoxidase staining and monoclonal antibodies against the virus specific antigens, in the mononuclear cells from the same blood specimens which were obtained for CMV pp65 monitoring. Altogether 322 blood specimens were analyzed. CMV disease was diagnosed in 12 (35%) patients during the first 3 months (first pp65 positive specimen mean 25 days, range 8-61 days) after transplantation. Concurrent HHV-6 antigenemia was detected in 10/12 (mean 14 days, range 6-22 days) and HHV-7 antigenemia in 9/12 patients (mean 25 days, range 10-89 days) after transplantation. HHV-6 usually appeared slightly before CMV. All CMV infections were successfully treated with ganciclovir and the CMV-antigenemia subsided. HHV-6 and -7 antigenemia also responded to the antiviral treatment, but more slowly than CMV. In conclusion, CMV infection was usually associated with HHV-6 and -7 antigenemia in liver transplant patients. The results support the suggestion that CMV, HHV-6 and -7 may have interactions. The clinical symptoms of CMV infection, may also be linked with HHV-6 or -7.

Stimulation of chicken lymphocytes by T- and B-cell mitogens.

Abstract Cultures of chicken spleen, peripheral blood, thymus, and bursal lymphocytes were tested for mitogenic stimulation by phytohaemagglutinin (PHA), concanavalin A (ConA), pokeweed mitogen (PWM), bacterial lipopolysaccharide (LPS), trypsin, and insulin. Spleen and blood leukocytes were stimulated by both the lectins and LPS, and also to some degree by trypsin and insulin as judged by increased incorporation of [ 3 H]thymidine into acid-insoluble material. This was observed in cultures incubated in serum-free medium as well as in the presence of foetal bovine serum or autologous plasma. Thymus cells were reproducibly stimulated by high concentrations of PHA. No significant responses were obtained in bursal cell cultures with any of the compounds tested. Removal of cotton wool-adherent cells from the spleen cell suspensions resulted in a subpopulation of cells which were stimulated by PHA but showed little response to ConA, PWM, or LPS. This procedure did not remove surface immunoglobulin-bearing cells from the original suspension. Both these enriched spleen lymphocytes and the unfractionated spleen, blood and thymus leukocyte cultures were effectively stimulated by a partially purified PHA but with a highly purified PHA preparation only at very high concentrations. These and other results suggest that the mitogenic components in crude PHA preparations are different for chicken and human or mouse cells.

Hepatitis A outbreak amongst intravenous amphetamine abusers in Finland

This article describes a widespread outbreak of hepatitis A virus (HAV) infection amongst drug abusers in Finland. Although attempts to demonstrate the virus in amphetamines failed, the infection was assumed to be linked to intravenous use of the drug. The unusual mode of transmission prompted us to analyse possible atypical clinical features as well as the spread of the virus to the general population, nowadays practically without protective immunity. Serologically verified cases that occurred in Helsinki were interviewed, their hospital records were analysed and their contacts were serology tested. Amphetamine lots, as well as faecal samples from patients, were examined with RT-PCR. Detailed information was obtained from 238 subjects, among whom 131 admitted drug abuse and 67 cases were classified as secondary cases. Phylogenetic analysis of virus strains from HAV-infected cases suggested a common origin, and epidemiological observations linked it with particular lots of amphetamine. Three cases died, and 3 presented with severe clinical disease. Icterus was more common among i.v. drug abusers than others. Infection with hepatitis A virus was probably related to the faecal contamination of amphetamine associated with the transportation of the drugs in the gastrointestinal tract.

Monitoring of Viral Load by Quantitative Plasma PCR during Active Cytomegalovirus Infection of Individual Liver Transplant Patients

ABSTRACT A quantitative PCR test, the Cobas Amplicor CMV Monitor, was used for the monitoring of viral load in the peripheral blood of 27 individual liver transplant patients and correlated with cytomegalovirus (CMV) pp65 antigenemia. Altogether, 243 specimens were analyzed. During the first 3 months, 20 patients showed PCR positivity which correlated with pp65 antigenemia. Of those, 13 patients developed symptomatic CMV infection 27 to 52 days after transplantation, with a significantly higher peak viral load in PCR and in pp65 assay compared with the seven asymptomatic infections (median 10,200 versus 2,240 copies/ml, P < 0.05, and median 100 versus 30 pp65-positive cells/50,000 leukocytes, P < 0.01). Five were primary infections of D+/R− cases (donor CMV seropositive and recipient seronegative) and demonstrated, except in one case, a high peak viral load (>10,000 copies/ml; range, 10,200 to 21,600 copies, and ≥50 positive cells, range, 50 to 800 cells). The peak viral loads of the six D+/R+ patients with symptomatic infection varied widely (range, 2,290 to 126,000 copies and 50 to 300 positive cells). Two D−/R+ patients developed symptomatic infection with a lower viral load (range, 1,120 to 6,510 copies and 25 to 100 positive cells). All symptomatic infections were successfully treated with ganciclovir. The asymptomatic infections all in D+/R+ patients with low copy numbers (<5,500 copies) were monitored until CMV disappeared. One of the seven PCR-negative patients had one sample with low antigenemia, but the subsequent specimens were all negative. The time-related correlation of the two methods was also good. In summary, quantitative PCR could equally well be used as the CMV pp65 assay for the monitoring of viral load in individual transplant patients.

Comparison of plasma polymerase chain reaction and pp65-antigenemia assay in the quantification of cytomegalovirus in liver and kidney transplant patients

BACKGROUNDnCytomegalovirus (CMV) is a significant problem in transplantation. The antiviral treatment is based on the clinical symptoms and the rapid laboratory diagnosis. Although polymerase chain reaction (PCR) methods have already been widely used, the clinical correlation of the findings is not clear.nnnOBJECTIVEnThe objective of this study was to investigate the usefulness of a quantitative plasma PCR test and compare it with the pp65-antigenemia test in the detection of clinically significant CMV infections in liver and kidney transplant patients.nnnSTUDY DESIGNnThe clinical material consisted of 253 consecutive blood samples was tested using a quantitative polymerase chain reaction test, Cobas Amplicor CMV Monitor (Roche) and pp65 antigenemia assay. Plasma was used for PCR and leucocytes were used for the antigenemia test.nnnRESULTSnCMV was detected in 89 out of 253 blood samples by one or both methods. PCR detected 78 (range 274-165000 copies/ml) and pp65 antigenemia test 79 (range 1-1500 positive cells/50000) of the positive findings. The sensitivity and specificity of PCR test was 86 and 94%, respectively. The PCR detected all clinically significant CMV infections (>10 positive cells in pp65 test) and infections which required antiviral treatment. In addition, the correlation between the two tests was almost linear.nnnCONCLUSIONSnThe quantitative PCR appears to be a suitable alternative to diagnose and monitor CMV infections in transplant patients.

Identification of antigenic components of Toxoplasma gondii by an immunoblotting technique

Proteins of Toxoplasma gondii were separated by SDS‐polyacrylamide gel electrophoresis with subsequent transfer to a nitrocellulose sheet by electrophoretic blotting. Immunologically reactive polypeptides were detected by human sera with previously known toxoplasma antibody levels. Heavy chain‐specific, peroxidase‐conjugated anti‐human immunoglobulins were used as the indicator antibodies for the separate identification of IgG and IgM reactive polypeptides. IgG toxoplasma antibodies reacted with several antigens of M r ≈27 000–67 000, while toxoplasma‐specific IgM seemed to detect only a few polypeptides. The M r of 35 000 for the dominating IgM reactive polypeptide was observed.

Improved sensitivity and specificity of enzyme immunoassays with P1-adhesin enriched antigen to detect acute Mycoplasma pneumoniae infection

An in-house P1-enriched (168-kDA protein) Mycoplasma pneumoniae antigen preparation was compared in IgG, IgA and IgM enzyme immunoassays (EIAs) to the respective EIAs employing crude antigen lysate, antigen prepared by Triton X-114 partition and two commercial antigens, one of which was an ether-extracted antigen and the other a P1-enriched antigen. In addition, three commercial kits from Sanofi Pasteur, Novum Diagnostica and Savyon Diagnostics were also assessed for comparison. Diagnostic sensitivity was studied with paired samples from adults (n=37) with acute respiratory illness interpreted as acute, recent or past infection to M. pneumoniae on the basis of the results of complement fixation test (CFT). If the consensus of at least two methods is taken as the true positive for acute infection, the diagnostic sensitivities of combined IgG and IgM EIAs were 100% for the Platelia(R), Sero MP and in-house EIAs whereas for the Novum EIAs and CFT- 97% and 74%, respectively. Moreover, the sensitivity of the P1-enriched antigen was proven superior on the basis of systematically highest OD(405 nm) ratios between convalescent and acute serum samples. Analytical specificity was studied by screening serum samples from 92 Finnish blood donors and 111 serum samples from cord blood. Diagnostic specificity was studied in a blind testing of 30 paired serum samples from infants with pneumonia of variable etiology. No single misinterpretation of acute infection from the group of samples with other respiratory diseases did occur. The present study confirmed and extended the earlier observations of the usefulness of P1-enriched antigen for reliable serologic diagnosis of acute M. pneumoniae infection.

CMV-induced class II antigen expression in various rat organs.

Abstracts Cytomegalovirus (CMV) is thought to trigger acute or chronic allograft rejection by inducing the expression of MHC class II antigens in the graft. This induction may be mediated by γ‐interferon or directly by CMV. In this study, we have investigated which structures in the rat kidney, liver, and heart are responsive to CMV‐induced class II expression in vivo. Rats were infected with rat CMV, the organs were harvested during the acute phase of infection, and the virus was demonstrated by culture from each organ. Direct CMV antigen detection was performed on frozen sections to demonstrate the detailed localization of CMV in the organs. In the kidney, CMV antigens were found in the vascular endothelium, in tubular cells, and scattered in the glomeruli. In the liver, the vascular structures and parenchyma contained CMV antigens. In the heart, CMV antigens were seen only in the capillary endothelium. Class II antigen expression was demonstrated by a monoclonal antibody and immunoperoxidase techniques. The induction of class II molecules was recorded in exactly the same cellular structures as those in which CMVantigens were detected.

Monoclonal antibody defining a human syncytiotrophoblastic polypeptide immunologically related to mammalian retrovirus structural protein p30

Antigenic material previously detected in human placental trophoblastic cells by immunoperoxidase staining using a goat antiserum against the feline RD114 retrovirus structural protein p30 was isolated by immunochromatography from normal syncytiotrophoblast. The antigen was used to immunize mice, and of the monoclonal antibodies produced by murine hybridomas an IgG1 was selected which reacted in enzyme immunoassay with the syncytiotrophoblast antigen and with purified RD114. This antibody, designated HPS-1, stained normal and neoplastic syncytiotrophoblasts in a manner similar to that of the goat antibodies, detected in immunoblotting a Mr = 130 000 polypeptide in cultured human choriocarcinoma cells and reacted in spot immunoblotting tests with purified preparations of mammalian retroviruses but not with an avian retrovirus. The polypeptide antigen may represent activation of human endogenous retroviral genes in syncytiotrophoblast.

High plasma HIV load in the CRF01-AE outbreak among injecting drug users in Finland.

An explosive outbreak of HIV-1 caused by the recombinant subtype AE (CRF01-AE) was detected in 1998 among Finnish injecting drug users (IDUs). These IDUs were compared with IDUs from the Amsterdam Cohort Study (ACS) infected with subtype B, to detect possible differences between 2 western IDU cohorts infected with different subtypes. Markers for progression (viral load and CD4+lymphocyte count) were compared between 93 IDUs with CRF01-AE and 63 IDUs with subtype B. Only persons with a seroconversion intervalu2005≤2 y were included. During 48 months of follow-up, both cohorts were similar in CD4+ cell decline, but the Finnish IDUs had 0.34–0.94 log10 copies/ml higher viral loads. The Amsterdam IDUs had a low viral load (<1000 copies/ml) significantly more often than the Finnish IDUs. The difference could not be explained by the use of antiretrovirals. The higher viral load may have contributed to the rapid spread of the recombinant virus in the Finnish outbreak.