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Biomedical subjects

G Jahn

Publications and source records attributed to G Jahn.

At least 55 records · Page 3Linked to original sources
PubMed1998

Transmission of cytomegalovirus to preterm infants through breast milk.

OBJECTIVE: To evaluate the rate of virus excretion through breast milk and the incidence and significance of postnatal cytomegalovirus (CMV) transmission from mothers to premature infants. DESIGN: Prospective study of mother-child pairs after preterm delivery before 32 weeks or birth weight < 1500 g. Exclusion of donor breast milk and of CMV-seropositive blood. Material used was maternal CMV serostatus, ear swab of the infants at birth, sequential screening of breast milk and children's urine. Methods used were CMV-DNA PCR and viral cultures on fibroblasts. RESULTS: During a 12-month period 56 mother-infant pairs with 67 preterm infants were studied. Twenty-seven women (48%) were CMV-seronegative at birth; breast milk samples and the infants' urine remained CMV-negative. Twenty-nine women were CMV IgG-seropositive; 23 of 27 seropositive breast-feeding mothers excreted CMV through milk (85%); 25 of 27 (93%) had CMV DNA-positive results. CMV infection occurred in 17 of 67 infants (25%). CMV transmission was exclusively found in infants of seropositive mothers who excreted CMV and breast-fed their infants; 17 of 29 exposed infants became infected (59%). In 12 patients (gestational age, 29.9 +/- 1.8 weeks) CMV was detected at a postnatal age beyond 8 weeks; 5 of these infants had mild signs of a viral infection. However, 5 extremely low birth weight infants (gestational age, 24.4 +/- 0.5 weeks) were infected at an age of 4 to 7 weeks; 4 of these infants had marked symptoms of an acute CMV infection. CONCLUSION: In mothers of preterm infants a high incidence of CMV excretion into breast milk was detected. There is evidence that the most immature infants are at the greatest risk to acquire an early and symptomatic CMV infection.

Cytomegalovirus Infections↗
PubMed1998

Evaluation of Murex CMV DNA hybrid capture assay for detection and quantitation of cytomegalovirus infection in patients following allogeneic stem cell transplantation.

Murex hybrid capture DNA assay (HCS) is a solution hybridization antibody capture assay for detection and quantitation of cytomegalovirus (CMV) DNA in leukocytes. To determine whether CMV HCS is sensitive enough to initiate and monitor antiviral therapy after allogeneic stem cell transplantation (SCT), 51 consecutive SCT recipients were prospectively screened for the appearance of CMV infection by HCS, PCR, and culture assays from blood samples. Preemptive antiviral therapy was initiated after the second positive PCR result in all patients, as previously reported, and HCS was not considered for clinical decision making. A total of 417 samples were analyzed. Of these, 21 samples were found to be positive by PCR and HCS, 88 samples were PCR positive but HCS negative, and 308 were negative by both assays. Concordance of results between PCR and HCS and between HCS and blood culture was observed in 78.9 and 95.9% of the samples assayed, respectively. PCR was found to be more sensitive than HCS, and HCS was more sensitive than the blood culture assay (P < 0.0001). Four patients with symptomatic CMV infection were PCR positive prior to the onset of CMV-related symptoms, whereas HCS detected CMV DNA in three patients prior to and one at onset of CMV disease. The numbers of genomes per milliliter of blood were higher in patients with symptomatic CMV infection than in those with asymptomatic CMV infection (P = 0.06). None of the HCS-negative patients developed CMV disease. Thus, all patients with CMV disease were correctly identified by HCS; however, the lower sensitivity limit of the HCS assay may still be insufficient to allow diagnosis of CMV infection early enough to prevent CMV disease in patients following allogeneic SCT.

Adolescent↗
PubMed1998

Human trophoblast cells are permissive to the complete replicative cycle of human cytomegalovirus.

Human trophoblast cells were permissively infected by human cytomegalovirus. The kinetics of viral immediate-early, early, and late gene expression was clearly delayed compared to that in fibroblasts. Productive infection was unequivocally proven by the detection of virion particles, infectious virus in trophoblast culture supernatant, and cell-to-cell spread of cytomegalovirus from infected trophoblasts to uninfected fibroblasts. These observations indicate that infected trophoblasts may be involved in maternofetal transmission of human cytomegalovirus.

Antigens, Viral↗
PubMed1998

Management of cytomegalovirus infection after solid-organ or stem-cell transplantation. Current guidelines and future prospects.

Recent developments in diagnosis and therapy of cytomegalovirus (CMV) infection have helped to reduce CMV-associated mortality following organ transplantation. However, CMV is still associated with significant morbidity in recipients of an allogeneic stem cell or solid-organ transplant. The clinical symptoms of active CMV infection per se and, most importantly, the prevalence of life-threatening CMV disease show broad variation between different patient populations depending on the type of transplant and the intensity of immuno-suppression. Therefore, management of CMV infection must be stratified according to risk profiles of a given patient population. In the past decade, novel diagnostic assays (such as rapid shell-vial culture, polymerase chain reaction, pp65 antigen assay and sensitive hybridisation techniques) have been developed. Broad variations in the ability of a given test to predict a positive or negative risk of developing CMV disease have been observed between different transplant modalities. Highly effective therapeutic agents against CMV, such as ganciclovir and foscarnet, have become available, improving the outcome of patients with CMV disease. Moreover, antiviral prophylaxis with ganciclovir or aciclovir has been shown to reduce CMV infection and CMV disease following organ transplantation. However, these drugs are often associated with considerable toxicity. Moreover, antiviral resistance to ganciclovir and foscarnet has been observed in recipients of organ transplants and, even more frequently, in patients with AIDS. Short courses of pre-emptive antiviral therapy, administered after CMV infection has been documented by sensitive diagnostic techniques prior to the development of clinical symptoms, help to reduce duration and incidence of adverse effects associated with antiviral drugs and are thus an attractive strategy compared with antiviral prophylaxis. Newer options, such as oral ganciclovir, cidofovir, benzimidavir (1263W94) and lobucavir, are currently under investigation and might further improve the management of CMV infection in recipients of solid-organ or stem-cell transplants.

Acyclovir↗
PubMed1997

Interstrain variation of immediate early DNA sequences and glycoprotein B genotypes in cytomegalovirus clinical isolates.

Cytomegalovirus (CMV) disease is associated with a high mortality in recipients of an allogeneic stem cell transplant. Apparent differences in biological behaviour have been noted among clinical CMV isolates. By amplifying specific functionally relevant regions of the CMV genome [immediate early (IE) exon 3, glycoprotein B (gB)], a possible association of strain variation and clinical symptoms of infection was analysed in 24 patients. A high number of genome mutations of the IE exon 3 region could be documented translating into amino acid changes of viral isolated of 8 out of 15 patients with symptomatic and 2 out of 9 patients with asymptomatic CMV infection. Identical IE mutations and gB types were observed in isolates from two different sites in 6 patients. gB strain 2 was found to be associated with symptomatic CMV infection (P = 0.03). Thus, apart from host factors viral factors might influence the virus-host interaction in severely immunosuppressed patients.

Amino Acid Sequence↗
PubMed1997

Semi-quantitative detection of cytomegalovirus DNA from native serum and plasma by nested PCR: influence of DNA extraction procedures.

The diagnostic implications of different procedures of DNA extraction were examined for the detection of HCMV DNA from sera and plasma of immunosuppressed patients. The detection limit of HCMV plasmid DNA from cell free seronegative plasma and serum by limiting dilution nested PCR decreased in the following sequence: phenol/chloroform > NaI-single tube method > proteinase K digestion equal to amplification of native sera and plasma. Nested PCR from native sera and plasma performed well and surpassed the proteinase K method in sensitivity for detection of serum DNAemia. Semi-quantitative determination of HCMV DNA titer present in native sera of immunosuppressed patients did not seem to be correlated to HCMV disease. When compared to the persistence of leukoDNAemia, the viral DNA titer in native plasma could only be observed in the acute phase of HCMV infection, an important phenomenon for diagnostic purposes. Correlation of serum DNAemia to virus culture revealed low positive and high negative predictive values. Predictive values of nested PCR from native sera for HCMV infection were not lower than those following organic DNA extraction. Despite its low correlation to viremia and virus isolation from any site, nested PCR from organic DNA extracts of serum or plasma is the most sensitive diagnostic tool of an ongoing HCMV infection. Additionally, semi-quantitative end point dilution nested PCR from native serum or plasma promises to be a rapid and easy tool for the monitoring of antiviral therapy.

AIDS-Related Opportunistic Infections↗
PubMed1997

Quantification of replication of clinical cytomegalovirus isolates in cultured endothelial cells and fibroblasts by a focus expansion assay.

A method for quantitative analysis of the growth properties of human cytomegalovirus (HCMV) in various cell culture systems was developed. Recent HCMV isolates are, in most cases cell associated, causing only limited cytopathic effect. This renders comparative analysis of the biological properties of such isolates difficult. The focus expansion assay described in this study is based on cocultivation of infected fibroblasts with a cell type of choice, relying on cell mediated infectivity. The extent of replication of a given isolate in cell culture is quantified by determining the size of resulting infectious foci. Analysis of various clinical isolates and laboratory strains indicated that this assay is a reliable and valid method to define growth properties of HCMV in cell culture. Remarkable differences in the cytopathogenicity of these isolates in fibroblasts as well as in endothelial cells were found. The assay will be useful in studies regarding cell tropism and virulence of recent HCMV isolates and for the quick and easy phenotypic characterization of HCMV deletion mutants.

Antigens, Viral↗
PubMed1997

Screening for CMV-specific T cell proliferation to identify patients at risk of developing late onset CMV disease.

Thirty patients undergoing allogeneic BMT were screened post-transplant together with their marrow donors for CMV-specific T cell proliferation and the occurrence of CMV disease. Twenty-one of these patients received a marrow transplant from an HLA-matched sibling donor, and nine from an HLA-matched unrelated donor. All these patients were either CMV seropositive and/or had received a transplant from a CMV-seropositive donor. Patients were monitored for CMV-viraemia until day +100 post-BMT by PCR and virus culture, and thereafter by virus culture only when clinically indicated. The proliferative T cell response was investigated at regular monthly intervals beginning on day +30. A proliferative response to HCMV (median, day +123) was documented in these patients between day +37 and +730 post-BMT. None of the patients with a documented CMV-specific T cell proliferation on day 120 (n = 17) developed CMV disease in the later post-transplant period, but of the patients lacking CMV-specific proliferation (n = 13), 30.8% developed CMV disease after day 120. Thus, patients lacking a CMV-specific T-helper cell response might benefit from sensitive screening for CMV infection and pre-emptive therapy after day +100.

Adolescent↗
PubMed1997

Cytomegalovirus monitoring by polymerase chain reaction of whole blood samples from patients undergoing autologous bone marrow or peripheral blood progenitor cell transplantation.

Sensitive screening for cytomegalovirus (CMV) by polymerase chain reaction (PCR) following autologous bone marrow or peripheral blood progenitor cell transplantation has not been evaluated. In a three-center study, 98 autograft transplant recipients were prospectively screened for CMV infection by PCR and culture techniques. At a median of 20 days (range, 3-28) after transplantation, 21 (39.6%) of 53 CMV-seronegative patients were PCR positive for CMV, and at a median of 17 days (range, 7-84) after transplantation, 19 (42.2%) of 45 CMV-seropositive patients were PCR positive for CMV. Low-level DNAemia (1-10 fg CMV DNA/mL blood) occurred for 1 week in 31 patients but was never associated with CMV disease. Of 9 patients who presented with at least two consecutive positive PCR results, 1 developed CMV pneumonia. No patients died because of CMV disease. Screening for CMV infection by PCR had a negative predictive value of 100% (as also observed after allogeneic transplantation), but its positive predictive value was significantly lower.

Adolescent↗
PubMed1996

The intracellular localization of human cytomegalovirus DNA in peripheral blood leukocytes during active infections by high-resolution fluorescence in situ hybridization.

Although viremia is an integral part of the pathogenesis of human cytomegalovirus (HCMV) disease, the interaction between HCMV and circulating leukocytes of actively infected patients remains an area of uncertainty. It is still a matter of dispute, whether leukocytes support viral replication with subsequent production of infectious virus. In a new approach we developed and applied a sensitive fluorescence in situ hybridization assay for the precise intracellular localization of HCMV genomes in leukocytes. It was shown that in vivo HCMV genomes were exclusively localized in the cytoplasm of leukocytes, indicating that the majority of these cells are virus carriers or abortively infected. Though this method easily detects single copy genes in metaphase chromosomes, the number of HCMV DNA positive leukocytes was significantly lower than the number of HCMV pp65 antigen positive cells. In relation to the pp65 antigen positive cells, only 1-4% of these cells were DNA positive. In addition, the much lower frequency of HCMV immediate early antigen positive leukocytes in comparison to the pp65 antigen positive cells and the impossibility of detecting other viral antigens support the hypothesis that the origin of pp65 found in leukocytes results mainly from protein uptake.

Antigens, Viral↗
PubMed1996

Fluorescence in situ hybridization with cosmid clones for the detection of human cytomegalovirus DNA in peripheral blood leukocytes.

Radioactive in situ hybridization techniques or enzymatic detection procedures of hapten-modified human cytomegalovirus (HCMV) probes have been widely used for studying the infection of peripheral blood leukocytes with HCMV. This report describes significant improvements in terms of signal resolution which can be obtained by applying a highly sensitive fluorescence in situ hybridization (FISH) technique in conjunction with a large subgenomic HCMV DNA probe. Three cosmid clones spanning 119.1 kb of the HCMV genome (230 kb) were used to construct the digoxigenin-11-dUTP-labeled probe which was found to be superior to a total HCMV probe representing the entire genome. Crucial hybridization parameters were analyzed systematically in order to ensure optimal resolution power and sensitivity. The protocol was successfully applied to HCMV-infected fibroblasts and peripheral blood leukocytes of 12 transplant patients and unambiguously facilitated the precise intracellular localization of HCMV genomes in infected cells. Because of its excellent resolution properties, accompanied by the virtual absence by any background staining, we recommend the use of this protocol as a sensitive approach for further virological analyses of the interactions between HCMV and peripheral blood leukocytes at the single-cell level.

Cloning, Molecular↗
PubMed1996

IgM-specific serodiagnosis of acute human cytomegalovirus infection using recombinant autologous fusion proteins.

Portions of three human cytomegalovirus (HCMV) polypeptides, which were shown previously to be highly reactive with patient sera, were expressed in Escherichia coli as autologous fusion proteins. Purified recombinant polypeptides were used as antigens in enzyme linked immunosorbent assay (ELISA) and compared against assays which use natural viral antigen from cell culture for their ability to improve IgM-specific serology of acute HCMV-infection. A fusion protein (CM2) which contained two copies of the C-terminal portion of pUL44 (p52, aa 297-433) and one copy of a highly reactive fragment of the major DNA-binding protein pUL57 (aa 545-601) proved to be superior in sensitivity and specificity compared to assays which use culture derived antigen. A construct expressing one copy of the fragments from pUL44 and pUL57 in fusion with the 54 amino terminal residues of pUL32 (pp150, aa 994-1048) did not lead to an improved sensitivity compared to CM2. Adversely, this polypeptide reacted with a number of sera from asymptomatic blood donors infected latently with HCMV indicating low specificity of this antigen for the detection of acute infection. Concordant results were obtained with an antigen that combined only the C-terminal portions of pUL44 and pUL32 (CM3). ELISA experiments with sequential sera from renal transplant recipients demonstrated that detection of IgM-antibodies using CM2 as antigen correlated closely with acute infection, whereas high levels of IgM-antibodies against CM1 and CM3 persisted for a month following acute HCMV-infection. These results indicate that the application of a single autologous fusion protein like CM2 as antigen for recombinant ELISAs can improve significantly IgM-serodiagnosis of acute HCMV infection.

Antibodies, Viral↗
PubMed1996

Cell types involved in replication and distribution of human cytomegalovirus.

As the number of patients suffering from severe HCMV infections has steadily increased, there is a growing need to understand the molecular mechanisms by which the virus causes disease. The factors that control infection at one time and the events leading to virus multiplication at another time are only beginning to be understood. The interaction of HCMV with different host cells is one key for elucidating these processes. Through modern techniques, much has been learned about the biology of HCMV infections in culture systems. In addition to endothelial cells, epithelial cells, and smooth muscle cells, fibroblasts are one cell population preferentially infected in solid tissues in vivo. From these sites of multiplication, the virus may be carried by peripheral monocytes and circulating endothelial cells to reach distant sites of the body. This would explain the multiorgan involvement in acute HCMV infection and the modes of viral transmission. From what has been learned mainly from human fibroblast culture systems, future studies will focus on how HCMV regulates the expression of its putative 200 genes in different host cells at different stages of cell differentiation and activation to result in viral latency and pathogenesis.

Acute Disease↗
PubMed1996

CMV infection after allogeneic bone marrow transplantation is associated with the occurrence of various autoantibodies and monoclonal gammopathies.

Recent findings indicate that the kinetics of B-cell reconstitution after marrow transplantation mimic normal ontogeny. The early B-cell repertoire during ontogeny is characterized by a high degree of autoreactivity and interconnectivity. Therefore, in a prospective analysis, 95 consecutive recipients of an allogeneic marrow transplant were screened for the occurrence of various autoantibodies and 47 of these 95 were also screened for monoclonal gammopathies. None of the patients developed antibodies specific for systemic autoimmune disorders. In contrast, a high prevalence of natural antibodies (79/95) was found early post-transplant, with 58 of these 79 patients developing two or more autoantibodies. According to multiple regression, the mean number of natural antibodies (95% confidence limits in parentheses) depends significantly (P = 0.006) on the status of CMV infection: 0.9 (0.4; 1.6) CMV-negative: 2.0 (1.0; 3.3) asymptomatic CMV infection; 3.1 (1.7; 5.0) CMV disease. Sex, age, underlying disease, conditioning therapy, acute graft-versus-host disease and CMV serology of donor and recipient pretransplant did not affect the number of natural autoantibodies. Monoclonal gammopathies were detected in 12/47 patients with a predominance of the IgG-kappa subtype. All these 12 patients suffered from a viral infection (CMV, n = 11: influenza strain A, n = 1). The high degree of self-reactivity post-transplant further supports the hypothesis that B-cell reconstitution mimics ontogeny. Moreover, these data indicate nonspecific polyclonal, CMV-mediated, presumably T-cell independent B-cell stimulation and disturbed T-cell regulatory function following allogeneic BMT.

Adolescent↗
PubMed1996

Tissue macrophages are infected by human cytomegalovirus in vivo.

On the basis of in vitro experiments, it has been suggested that cells of hematopoietic origin play a major role in the pathogenesis and latency of human cytomegalovirus (HCMV). To elucidate the in vivo importance of hematopoietic cells in acute HCMV infection, tissue sections from various infected organs were investigated by immunohistochemical double-labeling analyses. Monoclonal antibodies directed against distinct viral and cellular antigens were used to identify infected macrophages, polymorphonuclear cells, and lymphocytes. Macrophages and polymorphonuclear cells were targets for HCMV infection in different tissues. Viral proteins representing all stages of permissive HCMV infection were detected in macrophages, suggesting that these cells support the complete viral replication cycle. In polymorphonuclear cells, viral gene expression was restricted to the immediate early phase, indicating that these cells are abortively infected. These findings suggest that macrophages play an important role in the hematogenous spread of HCMV into solid organs.

Acute Disease↗
PubMed1996

Immunoglobulin A-specific serodiagnosis of acute human cytomegalovirus infection by using recombinant viral antigens.

The immunoglobulin A-specific reactivities of recombinant viral proteins from nine different reading frames of human cytomegalovirus were evaluated in enzyme-linked immunosorbent assay experiments. Antigen fragments of reading frames pUL32, pUL44, and pUL57 were identified as preferable antigens for immunoglobulin A serodiagnosis. Application of autologous fusion proteins which combine these polypeptides may be useful especially for the early detection of acute secondary human cytomegalovirus infection.

Acute Disease↗
PubMed1996

Human cytomegalovirus cell tropism and pathogenesis.

The human cytomegalovirus (HCMV) can cause lifelong infection with episodes of endogenous reactivation. Intrauterine fetal infection and infection of immunocompromised patients are well known to result in significant morbidity. Studies on HCMV cell tropism in vivo revealed three characteristics: (1) ubiquitously distributed cell types such as epithelial cells, endothelial cells and fibroblasts are the major targets of HCMV infection; (2) leukocytes circulating in the peripheral blood are susceptible to the virus, and (3) specialized parenchymal cells such as smooth muscle cells in the gastrointestinal tract and hepatocytes can also be infected. Questions to be resolved are, how the virus spreads throughout the organism, how it can impair the function of infected organs, and how it evades the host's immune response to establish lifelong infection. This chapter is focused on the role of HCMV-infected target cells for the pathogenesis of HCMV-associated disease in the acutely infected immunocompromised host.

Animals↗