Hierbij allereerst de Engelstalige bijdrage van Hanson:
Bron: forum Cort Johnson - XMRV-buzz.
Abstract: O_11
XMRV in Chronic Fatigue Syndrome: A Pilot Study
M.R. Hanson, L.L. Lee, L. Lin, D.E. Bell, D. Ruppert, D. S. Bell
Cornell University, Dept of Molecular Biology and Genetics, Ithaca NY, USA; State University of New York, Dept of Medical Anthropology, Buffalo NY, USA; Cornell University, School of Operations Research and Information Engineering, Ithaca NY, USA; State University of New York, dept of Pediatrics, Buffalo NY, USA
Background:
In October 2009, Lombardi et all repoted detection of XMRV in blood of persons with CFS. Since then, four studies that performed PCR analysis on DNA from blood of CFS patients failed to detect XMRV. The present blinded study was undertaken to determine whether XMRV could be detected in peripheral blood mononuclear cells (PBMCs) from three small groups of subjects from a single geographic area.
Materials and methods:
The 30 adult subjects of this study were divided into three groups of ten persons each: severely ill with CFS, recovered from CFS, and a control group lacking a CFS diagnosis at any time. Inclusion did not require that they became ill in the time frame or in the exact location of a cluster of CFS in Lyndonville, NY. All patients in group 1 ("severe CFS") currently meet Fukuda criteria. The individuals in group 2 ("recovered CFS") had met Fukuda criteria for CFS at one time in the past, but presently considered themselves recovered. One patient in group 1 and seven in group 2 are considered part of the "Lyndonville outbreak". The ten persons included in group 3 "healthy, non-household contact controls" have never experienced a CFS-like illness, are healthy, and have never lived with persons with CFS. Average ages of the three groups are 42.2, 39, 49.8, respectively; and M:F ratio is 2:8 6:4, 2:8. All thirty subjects completed seven different clinical survey instruments including the SF-36. Blood was collected in EDTA tubes and PBMCs and plasma separated within 24 hours of draw. Plasma from some samples was incubated with LNCaP cells. The Invitrogen Superscript (R) VILO (TM) cDNA Synthesis Kit was used to produce cDNA from RNA isolated from PBMCs or LNCaP cells. Nested PCR with USB Hot-Start IT FideliTaq was performed using the Gag O-Gag l primers originally described by Urisman et al. PCR products were separated on gels and any bands of approximately 400 bp were sequenced and analyzed by BLAST for similarity to the XMRV gag gene. PCR with mouse COX2 primers was performed on all cDNA preparations to rule out mouse cell contamination.
Results:
XMRV gag sequences were detected in eight of the "severe CFS", three of the "recovered CFS" and in one of the controls. Plasma from six blood samples, five CFS and one control, was incubated with LNCaP cells; the six cultures were passaged six times. The five cultures found to exhibit XMRV gag sequences were those inoculated with CFS patient plastma. Although group 2 members described themselves as recovered, their scores on the SF-36 were significantly lower than the healthy control group, according to Hoteling's T2 test. Tukey's multiple comparison of means indicates that there are highly significant differences between the scores of the "severe" and controls on all 7 instruments.
Conclusions:
XMRV gag sequences were detected in a higher percentage of severe or recovered CFS cases (55%) compared to controls (10%) with one-sided p-value=0.0118. Human plasma samples from CFS cases contain infectious XMRV. Our results corroborate those of Lombardi et al.
Ter info: samenvattingen XMRV-positieve studies conferentie
Moderator: Moderators
En vervolgens de studie van Pfost over XMRV bij kinderen. Te zijner tijd kan iemand dit abstract vertalen of samenvatten en dan wellicht weer verwijderen. Niet de bedoeling om dit net te vervuilen!
Nu is het nog even vers van de pers! Zelfde bron.
Abstract: P_19
Chronic Fatigue syndrome/ neuro immune diseases
Detection of infectious XMRV in the peripheral blood of children
M.A. Pfost1, K.S. Hagen1, F.W. Ruscetti2, J.A. Mikovits1
1Whittemore Peterson Institute, ARF, Reno, USA; 2NCI- Frederick, Laboratory of Experimental Immunology, Frederick, USA
Background: XMRV is a new human retroviral infection of as yet unknown pathogenic potential. Recent reports have found XMRV infection in 3% of healthy adult populations and high percentages in populations of immune compromised individuals and Chronic Fatigue Syndrome (CFS). The prevalence of XMRV infection has not been explored in families with CFS or in children. An understanding of the XMRV infection rate in children may be particularly helpful, given that 1 in 100 children in the US are diagnosed with neuroimmune disorders, including Autism Spectrum Disorder (ASD) and that CFS and childhood neuroimmune disorders share common clinical features including immune dysregulation, increased expression of pro-inflammatory cytokines and chemokines, and chronic active microbial infections. Thus, we hypothesized that XMRV infection may be detected not only in families with CFS but also in children with other neuroimmune disorders.
Methods: 66 subjects participated as family members of a parent or child diagnosed with a neuroimmune disease. Age, sex, date of onset, geographic location and length of illness were recorded. The study group consisted of 29 children, 2-18 years of age and 37 parents. 19 of the adults (51%) have a neuroimmune illness including CFS, fibromyalgia and Lyme disease and 17 of the children (59%) are diagnosed with ASD. One pair of 3 yr old twins have Niemann-
Pick type C, a neurodegenerative disease. 10 of the children (34%) were healthy siblings. Geographically, the subjects came from 11 states, 12% from the Southeast, 74% from the West, with 10% from NV, 8% from the Midwest and 6% from the Northeast. Sixteen families had more than one child participating including healthy siblings. Peripheral blood was drawn by a licensed phlebotomist under an approved IRB protocol, and shipped to the WPI for XMRV detection according to Lombardi et al. (Science, Oct 2009) including serology for antibodies to XMRV ENV, using PCR and RT-PCR on cultured PBMC nucleic acids as well as plasma isolation of XMRV to the LNCaP cell line. PCR products were sequenced at the Nevada Genomics Center using the ABI3730 DNA Analyzer.
Results: XMRV was detected in 55% of 66 cases of familial groups from 11 states. Sequencing of PCR products of env and gag confirmed XMRV. The age range of the infected children was 2-18. 17 of the children (including the identical twins) were positive for XMRV (58%) and 20 of the 37 parents (54%) were positive for XMRV. 14 of the 17 autistic children were positive for XMRV (82%). Of the 17 families, only one had all members of the family test negative for XMRV. In contrast, 16 of the families with neuroimmune disease, 9 families had at least 1 parent and child test positive for XMRV. 4 of the families had a parent test negative with a positive child, and 2 families had a parent test positive with the diseased child testing negative.
Conclusions: XMRV is observed in children with a wide spectrum of neuroimmune disorders and their family members. The significance of these findings is not clear.
No conflict of interest
Nu is het nog even vers van de pers! Zelfde bron.
Abstract: P_19
Chronic Fatigue syndrome/ neuro immune diseases
Detection of infectious XMRV in the peripheral blood of children
M.A. Pfost1, K.S. Hagen1, F.W. Ruscetti2, J.A. Mikovits1
1Whittemore Peterson Institute, ARF, Reno, USA; 2NCI- Frederick, Laboratory of Experimental Immunology, Frederick, USA
Background: XMRV is a new human retroviral infection of as yet unknown pathogenic potential. Recent reports have found XMRV infection in 3% of healthy adult populations and high percentages in populations of immune compromised individuals and Chronic Fatigue Syndrome (CFS). The prevalence of XMRV infection has not been explored in families with CFS or in children. An understanding of the XMRV infection rate in children may be particularly helpful, given that 1 in 100 children in the US are diagnosed with neuroimmune disorders, including Autism Spectrum Disorder (ASD) and that CFS and childhood neuroimmune disorders share common clinical features including immune dysregulation, increased expression of pro-inflammatory cytokines and chemokines, and chronic active microbial infections. Thus, we hypothesized that XMRV infection may be detected not only in families with CFS but also in children with other neuroimmune disorders.
Methods: 66 subjects participated as family members of a parent or child diagnosed with a neuroimmune disease. Age, sex, date of onset, geographic location and length of illness were recorded. The study group consisted of 29 children, 2-18 years of age and 37 parents. 19 of the adults (51%) have a neuroimmune illness including CFS, fibromyalgia and Lyme disease and 17 of the children (59%) are diagnosed with ASD. One pair of 3 yr old twins have Niemann-
Pick type C, a neurodegenerative disease. 10 of the children (34%) were healthy siblings. Geographically, the subjects came from 11 states, 12% from the Southeast, 74% from the West, with 10% from NV, 8% from the Midwest and 6% from the Northeast. Sixteen families had more than one child participating including healthy siblings. Peripheral blood was drawn by a licensed phlebotomist under an approved IRB protocol, and shipped to the WPI for XMRV detection according to Lombardi et al. (Science, Oct 2009) including serology for antibodies to XMRV ENV, using PCR and RT-PCR on cultured PBMC nucleic acids as well as plasma isolation of XMRV to the LNCaP cell line. PCR products were sequenced at the Nevada Genomics Center using the ABI3730 DNA Analyzer.
Results: XMRV was detected in 55% of 66 cases of familial groups from 11 states. Sequencing of PCR products of env and gag confirmed XMRV. The age range of the infected children was 2-18. 17 of the children (including the identical twins) were positive for XMRV (58%) and 20 of the 37 parents (54%) were positive for XMRV. 14 of the 17 autistic children were positive for XMRV (82%). Of the 17 families, only one had all members of the family test negative for XMRV. In contrast, 16 of the families with neuroimmune disease, 9 families had at least 1 parent and child test positive for XMRV. 4 of the families had a parent test negative with a positive child, and 2 families had a parent test positive with the diseased child testing negative.
Conclusions: XMRV is observed in children with a wide spectrum of neuroimmune disorders and their family members. The significance of these findings is not clear.
No conflict of interest
En nog eentje met CVS patienten. Wat nu nog volgens mij nog ontbreekt is de originele (positieve) bijdrage van 1. Lo, 2. Cheney en 3. ook van De Merleir die al (goeddeels) bekend zijn.
Detection of XMRV sequences in EBV-transformed B cell lines
J. Braneei J. camrrd, E. earcrai J. Areaf, B. creref, c. Cabrera
’lrsiCaixa Foundation, Retrowrology Lab, Barcelona, Urology Dept, Barcelona, Spain
Background: XMRV infection has been found in humans linked with prostate cancer and chronic fatigue syndrome (CFS). XMRV is able to infect a wide range of cells and has been found in a variety of cell types both in Vitro and in vivo, including B cells and other immune cell types.
Materials and methods: In order to determine the presence of XMRV sequences in B cells, we have screened 21 B cell lines available in our laboratory. These cells were generated by EBV immortalization of PBMC obtained from 11 CFS affected individuals (fulfilling both Fukuda and Canadian criteria), 5 healthy donors, 4 HIV infected individuals and 1 prostate cancer patient- DNA was extracted from dry cell pellets and XMRV sequences were amplified using a real-time PCR covering a 150bp pol sequence and using a nested approach in both gag and Env genes.
Results: Envelope amplification yielded positive bands in 4 out of 21 individuals tested, 3 CFS affected individuals and 1 healthy donor. However, gag amplihcation yielded only 3 positive samples (1 CFS affected individual, 1 healthy donor and one HlV+ patient). In contrast, Real-time PCR of Pol fragment detected 7 positives samples in 14 individuals tested (4 SFC , 2 donors and 1 HN+ individuals). To confirm the presence of XMRV sequences we performed sequence analyses of gag and env amplicons. The analysis of the three available gag sequences confirmed the XMRV characteristic 24-nt deletion, which is not found in any known exogenous MuLV. Sequences were 100% identical to reported XMRV sequences. Furthermore, envelope sequences
were also homologous to previously described XMRV sequences. ln this case, sequence variability was low or absent Interestingly, most of changes observed corresponded to G to A mutations that were accumulated in one positive sample.
Conclusions: Despite the discrepancies observed in the different PCR approaches using gag, pol or env sequences, our data suggest that EBV transformed B cell lines harbor XMRV
specific sequences, and therefore this cell type may represent a reservoir for XMRV contributing to its potential pathogenesis.
Detection of XMRV sequences in EBV-transformed B cell lines
J. Braneei J. camrrd, E. earcrai J. Areaf, B. creref, c. Cabrera
’lrsiCaixa Foundation, Retrowrology Lab, Barcelona, Urology Dept, Barcelona, Spain
Background: XMRV infection has been found in humans linked with prostate cancer and chronic fatigue syndrome (CFS). XMRV is able to infect a wide range of cells and has been found in a variety of cell types both in Vitro and in vivo, including B cells and other immune cell types.
Materials and methods: In order to determine the presence of XMRV sequences in B cells, we have screened 21 B cell lines available in our laboratory. These cells were generated by EBV immortalization of PBMC obtained from 11 CFS affected individuals (fulfilling both Fukuda and Canadian criteria), 5 healthy donors, 4 HIV infected individuals and 1 prostate cancer patient- DNA was extracted from dry cell pellets and XMRV sequences were amplified using a real-time PCR covering a 150bp pol sequence and using a nested approach in both gag and Env genes.
Results: Envelope amplification yielded positive bands in 4 out of 21 individuals tested, 3 CFS affected individuals and 1 healthy donor. However, gag amplihcation yielded only 3 positive samples (1 CFS affected individual, 1 healthy donor and one HlV+ patient). In contrast, Real-time PCR of Pol fragment detected 7 positives samples in 14 individuals tested (4 SFC , 2 donors and 1 HN+ individuals). To confirm the presence of XMRV sequences we performed sequence analyses of gag and env amplicons. The analysis of the three available gag sequences confirmed the XMRV characteristic 24-nt deletion, which is not found in any known exogenous MuLV. Sequences were 100% identical to reported XMRV sequences. Furthermore, envelope sequences
were also homologous to previously described XMRV sequences. ln this case, sequence variability was low or absent Interestingly, most of changes observed corresponded to G to A mutations that were accumulated in one positive sample.
Conclusions: Despite the discrepancies observed in the different PCR approaches using gag, pol or env sequences, our data suggest that EBV transformed B cell lines harbor XMRV
specific sequences, and therefore this cell type may represent a reservoir for XMRV contributing to its potential pathogenesis.