Clinically significant CMV (re)activation during or after radiotherapy/chemotherapy of the brain : Correlation with neurological deterioration and improvement upon antiviral treatment.

INTRODUCTION: For both patients with high-grade gliomas and multiple cerebral metastases, radio(chemo)therapy is the standard therapy. Neurological decline during treatment is rarely attributed to infections of the brain but to tumor progression or side effects of radiotherapy. CASE REPORTS: We present 4 cases of cytomegalovirus (CMV) viremia associated with neurological deterioration, which occurred during or shortly after radiotherapy and/or chemotherapy of the brain (brain metastases 2, high-grade glioma 1, carcinoma infiltrating brain 1). In all cases, neurological decline was sudden and unexpected, and causes such as increased intracranial pressure or tumor progression could be excluded radiologically. Treatment with dexamethasone and mannitol had no or only very short-term effects. General infections were either excluded or receding before the neurological symptoms occurred. All patients presented with decreasing levels of thrombocytes. In all cases, CMV (re)activation could be proven using blood test for CMV-DNA. The anti-CMV-IgG status suggested reactivation rather than a primary infection. One patient died within 72 h of onset of the symptoms (results of CMV tests were received postmortem). Diagnosis of 3 patients allowed successful administration of antiviral treatment, which greatly improved the general and neurological conditions of the patients within 48 h. DISCUSSION: Neurological deterioration during RT is hardly ever attributed to viral infections. These cases suggest that CMV reactivation and subsequent infection might actually be causative and has to be considered and treated. CONCLUSION: Further prospective studies verifying and investigating this observation in terms of frequency and clinical relevance seem indicated.

Performance characteristics of the VERSANT hepatitis C virus RNA 1.0 (kPCR) assay.

HCV RNA assays are of central importance for virological diagnostics and for clinical planning and monitoring of an antiviral combination treatment of chronic HCV infections. The objective of the pre-market evaluation of the VERSANT HCV RNA 1.0 Assay (kPCR) was to collect analytical performance data for this new method of HCV RNA quantification and to compare them with the high standards that exist in this context. The assay exhibited a specificity of 100%. The mean intra- and inter-assay imprecision was 14.1% and 14.6%, respectively. The detection limit was determined to be 16IU/ml (95% confidence interval: 11.9-30.6IU/ml) and consequently corresponded to the manufacturer's claims (i.e. 15IU/ml). The test exhibited linearity for all HCV genotypes in a broad range from 15 to 10(8)IU HCV RNA/ml. Hence, the kPCR assay in general is well suitable for HCV RNA determinations in clinical practice. However, in a methodological comparison, a considerable under-quantification of the concentrations of HCV genotype 2 and 3 isolates was detected. Provided that the assay's manufacturer will quickly remedy this shortcoming, the VERSANT HCV RNA 1.0 (kPCR) can be called a completely reliable technique for HCV RNA quantification in routine virological diagnostics.

[Changes in basic emotion recognition in patients with chronic low back pain. A cross-sectional study analyzing emotion recognition and alexithymia]. / Einschränkungen in der Erkennung von Basisemotionen bei Patienten mit chronischem Kreuzschmerz. Eine Querschnittstudie zur Analyse von Emotionserkennung und Alexithymie.

BACKGROUND: Persistent chronic pain leads to cortical changes in areas involved in the recognition of emotions. Wand et al. suggest a close correlation between the affective pain component and the extent to which these changes occur. As a consequence, the emotion profile may be influenced and difficulties in emotional communication may arise. MATERIALS AND METHODS: A total of 49 patients with chronic low back pain (CLBP) were classified as grade 1 + 2 or 3 + 4 using the Graded Chronic Pain Scale (GCPS) questionnaire. In all patients, the ability to recognize basic emotions coded through facial expression was assessed using the Facially Expressed Emotion Labeling (FEEL) test. Furthermore, the Toronto Alexithymia Scale-26 (TAS-26) was used to assess if the patients showed signs of alexithymia. Data of the GCPS grade 3 + 4 (n = 35) group were analyzed. RESULTS: Findings of the FEEL test indicate that the GCPS grade 3 + 4 group recognized the basic emotion 'surprise' significantly more often (p = 0.001) and showed a higher level of currently perceived anger than unaffected subjects. The TAS-26 showed that 28.5 % of the patients with CLBP were alexithym. CONCLUSION: The results suggest changes in the recognition of emotions and that patients with CLBP show signs of alexithymia. Further studies with larger sample sizes are required to confirm the detected trends.

Association of polymorphisms in solute carrier family 27, isoform A6 (SLC27A6) and fatty acid-binding protein-3 and fatty acid-binding protein-4 (FABP3 and FABP4) with fatty acid composition of bovine milk.

The main goal of this study was to develop tools for genetic selection of animals producing milk with a lower concentration of saturated fatty acids (SFA) and a higher concentration of unsaturated fatty acids (UFA). The reasons for changing milk fatty acid (FA) composition were to improve milk technological properties, such as for production of more spreadable butter, and milk nutritional value with respect to the potentially adverse effects of SFA on human health. We hypothesized that genetic polymorphisms in solute carrier family 27, isoform A6 (SLC27A6) fatty acid transport protein gene and fatty acid binding protein (FABP)-3 and FABP-4 (FABP3 and FABP4) would affect the selectivity of FA uptake into, and FA redistribution inside, mammary epithelial cells, resulting in altered FA composition of bovine milk. The objectives of our study were to discover genetic polymorphisms in SLC27A6, FABP3, and FABP4, and to test those polymorphisms for associations with milk FA composition. The results showed that after pairwise comparisons between SLC27A6 haplotypes for significantly associated traits, haplotype H3 was significantly associated with 1.37 weight percentage (wt%) lower SFA concentration, 0.091 lower SFA:UFA ratio, and 0.17 wt% lower lauric acid (12:0) concentration, but 1.37 wt% higher UFA and 1.24 wt% higher monounsaturated fatty acid (MUFA) concentrations compared with haplotype H1 during the first 3 mo of lactation. Pairwise comparisons between FABP4 haplotypes for significantly associated traits showed that haplotype H3 was significantly associated with 1.04 wt% lower SFA concentration, 0.079 lower SFA:UFA ratio, 0.15 wt% lower lauric acid (12:0), and 0.27 wt% lower myristic acid (14:0) concentrations, but 1.04 wt% higher UFA and 0.91 wt% higher MUFA concentrations compared with haplotype H1 during the first 3 mo of lactation. Percentages of genetic variance explained by H3 versus H1 haplotype substitutions for SLC27A6 and FABP4 ranged from 2.50 to 4.86% and from 4.91 to 7.22%, respectively. Tag single nucleotide polymorphisms were identified to distinguish haplotypes H3 of SLC27A6 and FABP4 from others encompassing each gene. We found no significant associations between FABP3 haplotypes and milk FA composition. In conclusion, polymorphisms in FABP4 and SLC27A6 can be used to select for cattle producing milk with lower concentrations of SFA and higher concentrations of UFA.

Sterol regulatory element binding transcription factor 1 (SREBF1) polymorphism and milk fatty acid composition.

Milk is known to contain high concentrations of saturated fatty acids-such as palmitic (16:0), myristic (14:0), and lauric (12:0) acids-that can raise plasma cholesterol in humans, making their presence in milk undesirable. The main objective of our candidate gene study was to develop genetic markers that can be used to improve the healthfulness of bovine milk. The sterol regulatory element binding transcription factor 1 (SREBF1) known to regulate the transcription of lipogenic genes together with SREBF chaperone and insulin induced gene 1 were the candidate genes. The results showed significant association of the overall SREBF1 haplotypes with milk production and variations in lauric (12:0) and myristic (14:0) acid concentrations in milk. Haplotype H1 of SREBF1 was the most desirable to improve milk healthfulness because it was significantly associated with lower lauric (12:0) and myristic (14:0) acid concentrations compared with haplotype H3 of SREBF1, and lower lauric acid (12:0) concentration compared with haplotype H2 of SREBF1. Haplotype H1 of SREBF1, however, was significantly associated with lower milk production compared with haplotype H3 of SREBF1. We did not detect any significant associations between genetic polymorphisms in insulin induced gene 1 (INSIG1) and SREBF chaperone and milk fatty acid composition. In conclusion, genetic polymorphisms in SREBF1 can be used to develop genetic tools for the selection of animals producing milk with healthier fatty acid composition.

No evidence of XMRV infection in immunocompromised patients and HIV-positive individuals from Germany.

BACKGROUND: Xenotropic murine leukaemia virus-related virus (XMRV) has been detected in patients with prostate cancer and chronic fatigue syndrome (CFS). The detection of XMRV in healthy individuals has raised concern about a possible virus transmission by blood products. However, recent studies challenge the association between XMRV and human disease. This study investigated whether or not XMRV is present in patients with altered immune function and individuals at increased risk of blood-borne viral infections in Germany. METHODS: We investigated 503 peripheral blood mononuclear cell (PBMC) samples from 240 patients with iatrogenic immune suppression (71 haematopoietic stem cell recipients, 132 solid organ transplant recipients, 37 others) and 311 PBMC samples from 302 patients with HIV-1 infection for the presence of proviral XMRV by real-time polymerase chain reaction (PCR). RESULTS: All 814 PBMC samples from 542 patients tested negative for XMRV DNA and positive for an internal herpesvirus saimiri (HVS) control. Human genomic DNA was detected in all samples, and 90% of the samples contained >10,000 cell equivalents per XMRV PCR reaction. CONCLUSIONS: Our failure to detect proviral XMRV provides evidence against the presence of XMRV in patients at increased risk of viral infections in Germany.

[HTLV-1-associated myelopathy/tropical spastic paraparesis: a differential diagnosis in multiple sclerosis]. / HTLV-1-assoziierte Myelopathie/tropische spastische Paraparese : Eine Differenzialdiagnose der Multiplen Sklerose.

Human T-cell lymphotropic virus 1 (HTLV-1) associated myelopathy/tropical spastic paraparesis (HAM/TSP) is a neurological disease caused by infection with HTLV-1. The disorder is very rare in Europe but endemic in many parts of the world. The pathogenesis is not clearly characterized but is based on a possibly immune-mediated injury of the cervicothoracic spinal cord. Clinically, HAM/TSP constitutes a slowly progressive spastic paraparesis associated with bladder dysfunction and often mimics the course of autoimmune and neurodegenerative diseases. The diagnosis is based on typical symptoms as well as detection of HTLV-1 specific antibodies and proviral HTLV-1 DNA or HTLV-1 RNA. The therapy is limited to symptomatic treatment. Transmission of HTLV-1 can occur vertically by breast feeding, through sexual contact or via infected blood products. Based on a clinical case report, we present here a current review on the pathophysiology, epidemiology, clinical manifestations, diagnosis and treatment of HAM/TSP.

[Prevention of nosocomial transmission of human immunodeficiency virus (HIV) from HIV-positive healthcare workers. Recommendations of the German Association for the Control of Viral Diseases (DVV) e.V. and the Society for Virology (GfV) e.V]. / Prävention der nosokomialen Übertragung von humanem Immunschwächevirus (HIV) durch HIV-positive Mitarbeiterinnen und Mitarbeiter im Gesundheitswesen. Empfehlungen der Deutschen Vereinigung zur Bekämpfung der Viruskrankheiten (DVV) e.V. und der Gesellschaft für Virologie (GfV) e.V.

To the best of our knowledge, the German Association for the Control of Viral Diseases (DVV) e.V. and the Society for Virology (GfV) e.V. are the first in Europe to provide precise recommendations for the management of health care workers (HCWs) who are infected with human immunodeficiency virus (HIV). Requirements for HIV-infected HCWs need to be clearly defined. With a permanent viral burden of less than or equal to 50 copies/mL, HIV-positive HCWs are allowed to perform any surgery and any invasive procedure, as long as the infected HCW uses double-gloving, undergoes follow-up routinely by occupational medicine professionals, undergoes a quarterly examination of viral burden, and has a regular medical examination by a physician who has expertise in the management of HIV. Unrestricted professional activity is only possible with a strict compliance to take antiretroviral therapy and if the HIV-infected HCW strictly adheres to the recommended infection control procedures. Complete compliance with the recommendation almost certainly leads to no HIV transmission risk in patient care.

Laboratory diagnosis of congenital CMV infection in newborns: Impact of pre-analytic factors.

BACKGROUND: To identify infants with congenital cytomegalovirus (cCMV) saliva polymerase chain reaction (PCR) is an ideal screening method. However, there are only few data on the influence of pre-analytic factors on the analytical sensitivity of the CMV PCR. OBJECTIVES: This study aimed to evaluate the performance of different swabbing materials, transport time and initial virus concentration regarding to the efficacy of recovery of CMV-DNA. STUDY DESIGN: Two CMV suspensions containing a high or low concentration of the laboratory strain AD 169 were prepared as test samples. Sampling was simulated by immersion of different swabs in these CMV suspensions and storing the swabs dry or in specified transport media. Transport conditions were modeled by storing the samples for defined time periods prior to DNA extraction and quantitative PCR analyses. Parallel analyses in two different laboratories allowed determination of lab to lab consistency. RESULTS: The duration of storage under the conditions analysed did not have a major effect on the recovery efficiency for the swabbing materials tested. With exception of flocked dry swabs, all tested swabbing materials demonstrated good recovery of CMV DNA. The flocked swab/eNAT system showed the best overall performance. CONCLUSIONS: All tested swabbing materials (with exception of the flocked dry swabs) seem to be well suited for recovery of CMV DNA and appropriate for use for the diagnosis of cCMV infection in symptomatic cases and in general cCMV screening programs of newborns.

Differences of nine drug resistance interpretation systems in predicting short-term therapy outcomes of treatment-experienced HIV-1 infected patients: a retrospective observational cohort study.

OBJECTIVE: Drug resistance interpretation systems are used to select the optimal antiretroviral therapy in HIV-infected patients. It is unclear how the systems perform in predicting therapy success and failure and in how far the interpretations are affected by insufficient drug levels. METHODS: The accuracy of nine different interpretation systems in predicting therapy outcomes was evaluated using virological, immunological, pharmacological, and clinical data of 130 patients treated at 13 outpatient centers. Individual susceptibility scores of the interpretation systems were converted into active drug scores (ADS) and correlated with therapy success and failure, defined as viral load reduction of equal to or more (n=66) and less than 1 log10 copies/ml (n=64) at three months after drug resistance testing. RESULTS: Three interpretation systems considered the respective therapies as more active compared to the other interpretation systems (p<0.01). These systems predicted therapy success better than the other systems, while the others performed better in predicting therapy failure. Thus, the overall rate of correctly predicted treatment outcomes was comparable between the different systems (73.1-80.0 %). Univariate and multivariate regression analysis revealed significant correlations between the ADS of all interpretation systems and virological therapy outcomes (p<0.0001). In contrast, only three interpretation systems were significantly correlated with immunological therapy outcomes in univariate and just one in multivariate models (p<0.05). Among 128 determinations of drug levels in 64 patient samples, 19.4 % revealed no detectable drug levels. The consideration of insufficient drug levels significantly improved the prediction accuracy of all interpretation systems (p<0.005). CONCLUSION: Differences between interpretation systems in predicting therapy failures and success need to be considered for future consensus algorithms. The prediction accuracy of interpretation systems can be improved by consideration of plasma drug levels.

Simple algorithm derived from a geno-/phenotypic database to predict HIV-1 protease inhibitor resistance.

BACKGROUND: Resistance against protease inhibitors (PI) can either be analysed genotypically or phenotypically. However, the interpretation of genotypic data is difficult, particularly for PI, because of the unknown contributions of several mutations to resistance and cross-resistance. OBJECTIVE: Development of an algorithm to predict PI phenotype from genotypic data. METHODS: Recombinant viruses containing patient-derived protease genes were analysed for sensitivity to indinavir, saquinavir, ritonavir and nelfinavir. Drug resistance-associated mutations were determined by direct sequencing. geno- and phenotypic data were compared for 119 samples from 97 HIV-1 infected patients. RESULTS: Samples with one or two mutations in the gene for the protease were phenotypically sensitive in 74.3%, whereas 83.6% of samples with five or more mutations were resistant against all PI tested. Some mutations (361, 63P, 71V/T, 771) were frequent both in sensitive and resistant samples, whereas others (241, 30N, 461/L, 48V, 54V, 82A/F/T/S, 84V, 90M) were predominantly present in resistant samples. Therefore, the presence or absence of a single drug resistance-associated mutation predicted phenotypic PI resistance with high sensitivity (96.5-100%) but low specificity (13.3-57.4%). A more specific algorithm was obtained by taking into account the total number of drug resistance-associated mutations in the gene for the protease and restricting these to certain key positions for the PI. The algorithm was subsequently validated by analysis of 72 independent samples. CONCLUSION: With an optimized algorithm, phenotypic PI resistance can be predicted by viral genotype with good sensitivity (89.1-93.0%) and specificity (82.6-93.3%). The reliability and relevance of this algorithm should be further evaluated in clinical practice.

Phenotypic HIV-1 resistance correlates with treatment outcome of nelfinavir salvage therapy.

In order to analyse whether drug sensitivity testing would be beneficial for clinical decision-making in heavily pretreated patients, we retrospectively studied viral genotype and phenotypic drug resistance in 12 HIV-1-infected patients, each of them with a history of failing at least one therapeutic regimen including one or two protease inhibitors (PIs). The salvage therapy included nelfinavir as new PI in all cases. Four patients showed a sustained and five patients a transient viral load decrease. Three patients failed to show a significant decline of plasma HIV-1 RNA. In the baseline samples of these cases, resistance against all components of their combination therapy could be detected, whereas at least one antiretroviral drug was still active in the cases with transient treatment response. All patients with sustained therapy response harboured viruses that were either fully sensitive or resistant to only one of the drugs administered. In our study, phenotypic drug resistance was predictive for the success of antiretroviral salvage regimens.

Decreased alloreactivity after vaccination against hepatitis B.

The beneficial effect of blood transfusions on renal allografts disappeared at about the same time when hepatitis B vaccination was introduced in dialysis patients. To determine possible immunosuppressive effects of hepatitis B vaccination, we studied alloreactivity during the course of immunization. Fourteen healthy subjects without serological evidence of hepatitis B were routinely immunized against hepatitis B surface antigen. Plasma and mononuclear cells were isolated and frozen before immunization and after vaccination, respectively. Mean alloreactivity measured by [3H]thymidine uptake decreased from 64,772 cpm before immunization to 40,213 cpm after the third immunization. In crossover experiments, cellular modulation and plasma-dependent modulation of alloreactivity were observed. The immunosuppressive effect of plasma taken 4 weeks after the third vaccination correlated (r = 0.9) highly significantly (P < 0.005) with the anti-hepatitis B surface antigen antibody titer. Taken together, our data strongly suggest that hepatitis B vaccination is capable of reducing allogenic reactivity.

Conversion of recurrent delta-positive hepatitis B infection to seronegativity with famciclovir after liver transplantation.

BACKGROUND: Recurrent hepatitis B infection after liver transplantation is associated with poor graft and patient survival. Famciclovir is a nucleoside with virostatic action in hepatitis B infection. We report the case of a 51-year-old patient who developed recurrent delta-positive hepatitis B infection after liver transplantation. After famciclovir treatment, he became seronegative for hepatitis B early and hepatitis B surface antigens and developed protective anti-hepatitis B surface antibody titers. METHODS: After recurrent hepatitis B was confirmed, treatment with famciclovir was initiated. RESULTS: Eighteen days after starting famciclovir, the patient became seronegative for hepatitis B early antigen and delta antigen, and hepatitis B virus DNA was no longer detectable in serum. Three months later, the patient became hepatitis B surface antigen negative and remains well 16 months later with increasing anti-hepatitis B surface levels. CONCLUSIONS: Antiviral treatment with famciclovir may be useful in treatment of delta-positive hepatitis B infection following liver transplantation. Further evaluation of famciclovir in treatment and prevention of hepatitis B in these patients is warranted.

Rapid, phenotypic HIV-1 drug sensitivity assay for protease and reverse transcriptase inhibitors.

BACKGROUND: Development of drug resistance is one of the major reasons for the failure of antiretroviral therapy of HIV-1 infection. Knowing the drug sensitivity-resistance profile of viruses present in a patient prior to treatment or change in treatment could help to optimize therapy. OBJECTIVE: Development of a rapid standardized phenotypic HIV-1 drug sensitivity assay for protease (PR) and reverse transcriptase (RT) inhibitors. DESIGN: The PR gene (codons 1-99) and the 5' part of the RT gene (codons 1-300) of HIV-1 is amplified from the plasma of infected individuals by RT-PCR and ligated into a proviral clone of HIV-1 containing a deletion of the PR gene and the 5' part of the RT gene. Bacteria are transformed with the ligation product and plasmid DNA is prepared from a library of transformed bacteria. The plasmid DNA is transfected into 293 T cells and recombinant virus is harvested from the supernatant of the transfected cells 2 days after transfection. The sensitivity of the recombinant virus is determined with the help of a sensitive indicator cell line. RESULTS: Recombinant viruses were generated with high efficiency. Determination of the drug sensitivity of the recombinant viruses with an indicator cell line was highly reproducible. The recombinant viruses accurately reflected the sensitivity-resistance profile of the parental viruses. The phenotypic drug sensitivity determined by this assay correlated well with the treatment history of patients. CONCLUSION: This assay system should allow rapid, high-throughput analyses of phenotypic HIV-1 drug sensitivity for PR and RT inhibitors. Due to the efficient generation of recombinant viruses, propagation of the recombinant viruses in cell culture is not required prior to the determination of the sensitivity of the recombinant viruses. The risk of selecting fitter non-resistant viruses due to culture conditions is minimized.

Early diagnosis and successful treatment of acute cytomegalovirus encephalitis in a renal transplant recipient.

We report the case of a 40-year-old male HIV-negative renal transplant patient with allograft rejection and immunosuppressive therapy who presented with acute cytomegalovirus (CMV) encephalitis. CT and MRI of the brain were normal but EEG showed diffuse slowing and dysrhythmia. In cerebrospinal fluid (CSF) initially 81 cells/microliters were found and immunocytochemistry showed a decreased CD4/CD8 ratio and increased values of activated lymphocytes, natural killer cells and immunoglobulin-containing cells. CMV-specific IgM antibodies in CSF and serum, immunostaining of CMV antigen in CSF cells and virus culture from CSF and urine were negative. During the first 3 weeks of illness no intrathecal production of immunoglobulins could be detected. Early diagnosis of CMV encephalitis was made by in situ hybridization (ISH) on CSF cell preparations and the polymerase chain reaction (PCR) which was positive in CSF and blood. On day 26 diagnosis was confirmed by detection of CMV-specific intrathecal IgG production. The patient was treated with ganciclovir, anti-CMV immunoglobulins and intrathecal beta interferon. He recovered completely after 2 months. Our data demonstrate the usefulness of ISH and PCR in the early diagnosis of CMV encephalitis and perhaps may encourage the use of intrathecal beta interferon in other patients with this disease.

Comparative analysis of intrathecal antibody synthesis and DNA amplification for the diagnosis of cytomegalovirus infection of the central nervous system in AIDS patients.

We evaluated 49 paired cerebrospinal fluid (CSF) and serum samples of 35 patients infected with the human immunodeficiency virus type 1 (HIV-1) for laboratory evidence of cytomegalovirus (CMV) infection. The patients were grouped according to clinical criteria as probable CMV encephalitis/polyradiculomyelitis, CMV retinitis, cerebral toxoplasmosis, progressive multifocal leukoencephalopathy, HIV-1-related cognitive/motor complex, HIV-1-associated myelopathy, and other neurological diseases. Paired CSF and serum samples were analysed for CMV deoxyribonucleic acid (DNA) by polymerase chain reaction (PCR), quantitative intrathecal synthesis of immunoglobulin G (IgG) antibodies specific for recombinant phosphoprotein 150 (pp150) of CMV and CMV-specific serum IgM. Intrathecal synthesis of pp150-specific IgG was detected in 26% of patients (9/35), serum IgM was found in 23% of patients (8/35), and PCR of CSF was positive in 11% of patients (4/35). Detection of CMV-specific DNA in CSF preceded the intrathecal antibody synthesis in three patients for whom serial samples were available. PCR results of the CSF became negative in one patient with CMV polyradiculomyelitis after successful therapy with 9-[2-hydroxy-1-(hydroxymethyl) ethoxymethyl] guanine (DHPG). PCR has a higher diagnostic specificity in the acute phase of CMV infection than intrathecal antibody synthesis. The serum IgM response to CMV cannot be used to monitor a compartmentalized immune response in the central nervous system while an intrathecal immune response seems to be associated with recovery either spontaneously or as a result of treatment.