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Abstract: Background: Herpes virus infections are suspected to be involved in the pathogenesis of atherosclerosis. Objective and method: Viral DNA of herpes simplex virus (HSV), Epstein-Barr virus (EBV) and cytomegalovirus (CMV) was analyzed by real-time PCR on 48 biopsies from atherosclerotic plaques extracted by end-arterectomy (46 coronary arteries, 2 carotid arteries), and in tissue from non-atherosclerosis vessels from the same patient as controls (23 internal mammary arteries, 43 saphenous veins). Results: HSV-1 DNA was detected significantly more frequently in plaques (35%) than in control veins (9%, P = 0.006). However, the frequency of HSV-1 DNA detection in the internal mammary artery grafts was as high as in plaques (22%, P = 0.28). CMV and EBV DNA were exclusively found in plaques but not in controls, with 10% for CMV (P = 0.06 versus veins, P = 0.17 versus graft arteries) and 2% for EBV (P = 1.0), respectively. HSV-2 was neither detected in plaques nor in controls. Herpes viral DNA was significantly associated only with arterial hypertension but not with other classical risk factors (P = 0.02), in accordance with the hypothesis that herpes viral infection may alter the vessel wall. Conclusion: We conclude that herpes viral infections may have a role in atherosclerosis and that the presence of herpes viral DNA in the grafts used for bypass surgery might constitute a potential risk for atherosclerosis or restenosis. [Copyright &y& Elsevier]

Background: Tick borne encephalitis virus (TBEV), is a human flavivirus causing tick borne encephalitis (TBE), a viral infection of the central nervous system endemic in Europe and Asia. Objectives: To develop a reverse transcription polymerase chain reaction (RT-PCR) assay based on quantitative real-time RT-PCR technology (TaqMan) for detection and quantification of TBEV RNA. The test includes an internal control (IC) to avoid false negative results. Study design: The system was established and validated using wild-type (WT) non-infectious synthetic RNA representing a fragment of the 3′ non-coding region of the TBEV genome. In addition, synthetic RNA differing from the WT synthetic RNA by a unique probe binding region was used as IC to monitor the overall efficiency of the RT-PCR. Results: The analytical sensitivity of the assay was at least ten copies of the TBEV synthetic transcript in presence of 50 copies of the IC. Successful amplification was obtained for different strains within the TBEV complex (Hypr, Hochosterwitz, Laibach, Elsass=Alsace, ZZ9, Wladiwostok). Among 14 serum and 21 cerebrospinal fluid (CSF) samples obtained from 28 patients with clinical suspicion of TBEV 1 CSF sample tested positive for TBEV RNA. In addition, no TBEV RNA could be detected in blood samples obtained from three vaccinated people 1 and 3 days post-vaccination. Thus indicating that a positive result is unlikely to be caused by recent vaccination. Conclusions: A quantitative, highly sensitive and specific real-time RT-PCR assay has been developed for the detection of TBEV RNA. Inclusion of an IC is important to monitor the possible occurrence of false-negative results caused by the presence of inhibitory factors. This assay should be an important asset for the routine laboratory detection of TBEV RNA. [Copyright &y& Elsevier]

Background: The placental and umbilical blood vessel endothelia may play an important role in human cytomegalovirus (CMV) transmission and viral propagation in the fetus. Objectives: We propose that CMV infection promotes endothelial cell viability for virus replication. Furthermore, we suggest that certain viral mechanisms are established to delay or inhibit the cellular response to undergo apoptosis. Study design: We have established a model of CMV infection in primary endothelial cells (HUVECs) in which productive infection with the endothelial-adapted strain of CMV, strain 4010, inhibits apoptosis induced by growth factor withdrawal. Apoptosis was measured by assessment of nuclear changes by TUNEL staining. Results: Our results indicate that CMV infection at 48–96 h significantly protects the endothelial cell from apoptosis induced by growth factor withdrawal. We found that uninfected cells in the CMV-infected cell cultures, but not CMV-infected cells, were susceptible to apoptosis during growth factor withdrawal. Our studies examine the pro-apoptotic pathways that are inhibited and the anti-apoptotic pathways that are activated during CMV infection. The pro-apoptotic protein, caspase-3, is activated during growth factor withdrawal in mock-infected HUVECs, but activation is reduced in CMV-infected HUVECs. Conclusions: During infection, the CMV-encoded chemokine receptor US28 is expressed on the infected cell surface. Previously, we have shown that stimulation of US28 with the chemokine RANTES activates cellular proliferative responses in endothelial cells. It is likely, therefore, that US28 might activate anti-apoptotic mechanisms during CMV infection. [Copyright &y& Elsevier]

Background: Multiple epidemiological studies have suggested that cytomegalovirus (CMV) infection is associated with atherosclerotic disease. However, conclusive proof that the virus is directly related to the progression of the disease is still lacking. Objectives: The goal of this study was to investigate whether MCMV is able to exacerbate the atherosclerotic process in atherosclerosis-susceptible mice. Study design: apoE knockout mice kept on a chow diet were sacrificed at both 2 and 20 weeks post infection (p.i.). C57Bl/6J mice fed an atherogenic diet were sacrificed at 2 weeks p.i. Lesion area, lesion composition (endothelial cells and smooth muscle cells) and inflammatory influx (T-lymphocytes and macrophages) in lesions were determined. The former one was determined by means of a microscope coupled to a computer-assisted morphometry system. The latter ones were scored after immunohistochemical staining. Results: In the chronic phase of the infection mean lesion size was significantly increased after MCMV infection in the apoE knockout mice. This increase could to a large extent be attributed to a significant increase in type V lesion area after MCMV infection. Also, a significant increase in T-lymphocyte influx was observed in the acute phase of the infection in lesions from apoE knockout mice after MCMV infection while this effect was absent in C57Bl/6J mice. After MCMV infection no increase was observed in macrophage, smooth muscle cell and endothelial cell number in lesions from both mice strains. Conclusions: MCMV infection may exacerbate the atherosclerotic process in apoE knockout mice by means of an acute lymphocytic inflammatory response. In contrast to the MCMV induced effect in apoE knockout mice, MCMV infection did not increase the influx of T-lymphocytes in atherosclerotic lesions of C57Bl/6J mice. [Copyright &y& Elsevier]

Background: Cytomegalovirus (CMV) infections are very common in the general population. Clinical CMV disease, particularly CMV pneumonitis, greatly impacts the morbidity and mortality of immunosuppressed patients. Objective: To present an overview of the basic aspects of the biology, epidemiology, and clinical features of CMV in relation to the available diagnostic and therapeutic approaches in adult patients. Methods: Review of the medical literature on cytomegalovirus infection and disease in adult hosts, with a focus on approaches to diagnosis and treatment of CMV respiratory disease in immunosuppressed hosts. Conclusions: Cytomegalovirus infections are likely to remain a significant cause of morbidity and mortality among immunosuppressed patients. Important aspects of the biological events underlying the transition from infection to clinical disease remain unclear. Despite that, considerable progress has been made in the design of improved diagnostic techniques and the development of antiviral agents. Preventive and particularly preemptive therapeutic approaches demand further technical improvements in diagnostic testing. At present, the emphasis in the search for improved diagnostic testing rests on the development of quantitative methods for early detection of the increased viral replicative activity that presumably precedes the onset of CMV disease in infected individuals. [Copyright &y& Elsevier]

Background: Human herpes virus-6 (HHV-6) was first isolated in 1986. It has been shown to cause exanthema subitum and has been associated with various other diseases. HHV-6 infection is widespread, and more than 90% of the population have antibodies against HHV-6 at the age of 2 years. Once acquired, the virus remains latent in the body. This makes it difficult to draw any conclusions about a causal relationship between the demonstration of HHV-6 and a specific disease. Objectives: This work was to develop a μ-capture HHV-6 IgM enzyme linked immuno sorbent assay (ELISA) for use in routine diagnosis and for wide scale patient population analysis. Study design: A μ-capture HHV-6 IgM ELISA was established. A total of 682 sera consisting of 585 sera from Danish blood donors and 97 sera from patients with autoimmune antibodies were analysed in the HHV-6 IGM ELISA. One hundred and ninety-two sera had earlier been analysed for total HHV-6 antibody content in a competitive ELISA, 94 sera were analysed for cytomegalovirus (CMV) IgM and 57 sera for Epstein Barr virus (EBV) antibodies, using different ELISA assays. The results for 12 primary infections with HHV-6 are also reported. Results: A HHV-6 IgM optical density (OD)-ratio was calculated according to a constant positive control. An empirical cut off of 0.5 HHV-6 IgM OD-ratio was chosen (with regard to the 10 HHV-6 seroconverters), which resulted in a specificity of 97.5% of the HHV-6 IgM ELISA. Two of the three donor sera with HHV-6 IgM OD-ratios more than 1.05 had total HHV-6 antibody titers significantly above the group with IgM OD-ratios below 0.7 consisting with HHV-6 reactivation. There was no cross reactions to EBV or CMV IgM positive sera. Conclusion: The HHV-6 IgM ELISA seems valid to diagnose primary HHV-6 infection in particular in combination with the HHV-6 total antibody assay. [Copyright &y& Elsevier]

Background: Human herpesviruses may cause infections of the central nervous system (CNS). The early diagnosis of herpesvirus-associated neurological diseases is of high importance. Objectives: The objective of this paper is to summarize the experience gained with the diagnosis of herpesvirus infections of the CNS at our institute by polymerase chain reaction (PCR)-based assays within the past few years. Study design: A retrospective analysis of herpesvirus desoxy ribonucleic acid (DNA)-positive cerebrospinal fluid (CSF) samples was performed, with particular emphasis on data obtained by quantification of virus DNA in CSF by newly established real-time quantitative PCR assays. Results: Herpesviruses were found in 26.6% of all virus-positive CSF samples detected at our institute between 1995 and 2001. The overall broad testing for different herpesviruses from CSF has led to an increase in the detection rate, especially in relation to varicella zoster virus (VZV)-associated CNS disease. The herpesvirus DNA load in CSF was investigated by TaqMan real-time PCR assays that were established for the individual herpesviruses. The amount of virus varied among the individual diseases, associated with herpes simplex virus type 1, herpes simplex virus type 2, VZV and cytomegalovirus, while for Epstein-Barr virus and human herpesvirus type 6 only low levels of virus were detectable in CSF. Conclusions: A generally broad testing for different herpesviruses in CSF samples is highly recommended. In addition, determination of the virus DNA level in CSF by quantitative assays seems to be of high importance for elucidating aspects concerning the prognosis of disease, the prediction of distinct CNS manifestations, and possibly the differentiation between specific virus-associated disease and unspecific presence of virus in CSF, especially in immunocompromised patients. [ABSTRACT FROM AUTHOR]

Background: The viruses of the Herpesviridae family, in particular herpes simplex virus types 1 (HSV-1) and 2 (HSV-2), cytomegalovirus (CMV), Epstein–Barr virus (EBV), varicella zoster virus (VZV), and human herpesvirus 6 (HHV-6), are responsible for numerous infections of the central nervous system (CNS). These infections manifest as diverse clinical signs, many of which are not specific. The diagnosis of these infections is necessary to make it possible to adapt treatment appropriately, as treatment is specific for the particular virus concerned. Objectives: To apply a polymerase chain reaction (PCR) technique for the diagnosis in a single reaction of the six herpesviruses most frequently detected in the cerebrospinal fluid (CSF) and to analyse clinical events in patients presenting positive results in PCR for herpesviruses. Study design: We studied 141 patients, from whom 180 CSF samples were collected. The clinical files of the patients were consulted retrospectively, and a list of clinical signs was recorded. After testing by targeted PCR, at the clinician''s demand, we tested these samples by herpes consensus PCR, which detects six herpesviruses (HSV-1, HSV-2, CMV, EBV, VZV, HHV-6), in a single PCR. Results: Targeted PCR tests identified 25 CSF samples (13.9%), corresponding to 18 patients (12%), as positive. The herpes consensus PCR test detected 49 samples (27.2%) as positive, resulting in the identification of 54 individual viruses (four samples displayed co-infection) from 39 patients (27%). 130 CSF samples, from 101 patients, tested negative by both techniques. 23 HIV-positive patients (30.6%), three HIV-negative immunocompromised patients (27%), and 14 immunocompetent patients (25%) were CSF PCR-positive. In HIV-positive patients, CMV was the virus most frequently identified (13%), followed by EBV (10.6%), VZV (5.3%) and finally HSV-1 and HSV-2 (both 1.3%). We did not detect HHV-6 in any of these samples. We detected only HSV-2, EBV and VZV in the 11 HIV-negative immunocompromised patients. CSF samples of immunocompetent patients contained mostly VZV (9%) and HSV-1 (7.3%). Conclusions: The herpes consensus PCR for a given virus was more sensitive than the standard, targeted PCR used in our laboratory. The clinical signs presented by patients infected with HSV-1, HSV-2 and CMV were similar to those reported in previous studies. For VZV, we report the possibility of mild, transient cerebral viral reactivation. Our data on the detection of EBV by PCR suggest that the PCR test is of predictive value for cerebral lymphoma in immunocompromised patients. The possible role of HHV-6 in a subacute neurological disorder merits further investigation. [ABSTRACT FROM AUTHOR]

Background: Herpesviruses may be associated with various types of central nervous system (CNS) infections. Herpes simplex encephalitis (HSE) has to be considered one of the most severe diseases. As effective antiviral drugs are available, rapid and reliable diagnosis has become important. Objectives: To describe polymerase chain reaction (PCR) and serological methods for the detection of herpesvirus-induced CNS infections by the example of HSE. Study design: 620 cerebrospinal fluid (CSF) and 2400 serum samples from 2700 selected hospitalized patients with clinical suspicion of encephalitis were tested for herpes simplex virus (HSV) as well as varicella-zoster virus (VZV) DNA and HSV-specific antibodies, respectively. Results: HSV-1 DNA could be detected in eight and HSV-2 in three patients with focal encephalitis. In addition, HSV-2 DNA was found in two newborns with encephalitis and two adults suffered from transverse lumbar myelitis. One VZV DNA-positive patient had developed herpes zoster accompanied by meningoencephalitis, and in the other an encephalitis without cutaneous rash was diagnosed. Intrathecal antibody synthesis could be measured when CSF was cleared from viral DNA. Conclusions: The detection of viral DNA by PCR technique has become the “gold standard” method for laboratory diagnosis of herpesvirus infections of CNS. Serodiagnosis may be useful to confirm the diagnosis retrospectively. [ABSTRACT FROM AUTHOR]