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Reverse Line Blot (reverse + line_blot)
Selected AbstractsLow-risk HLA genotype in Type 1 diabetes is associated with less destruction of pancreatic B-cells 12 months after diagnosisDIABETIC MEDICINE, Issue 12 2007M. Spoletini Abstract Aims The role of human leukocyte antigen (HLA) genes in the susceptibility to Type 1 diabetes (T1DM) is well known. However, we do not know whether the degree of pancreatic B-cell destruction depends on different HLA genetic risk. The aim of this study was to analyse the influence of DRB1* and DQB1* genes on the rate of pancreatic B-cell loss in a prospective series of 120 consecutive newly diagnosed T1DM subjects in the first 12 months after diagnosis. Methods Patients were typed for HLA-DRB1* and DQB1* loci by a reverse line blot assay using an array of immobilized sequence-specific oligonucleotide probes. C-peptide, insulin requirement and glycated haemoglobin (HbA1c) were determined at diagnosis and every 3 months for 12 months. The variance of C-peptide as evidence of B-cell loss during follow-up was analysed using the general linear model for repeated-measures procedure. Results Fasting C-peptide in T1DM subjects with low HLA genetic risk was significantly higher when compared with subjects with moderate or high HLA genetic risk from time of diagnosis up to 12 months (P = 0.007 and P = 0.0002, respectively). Nonetheless, the changes in C-peptide levels over a 12-month period did not differ significantly between T1DM subjects with different HLA genetic risks. Conclusions Low-risk HLA genotype in T1DM is associated with less destruction of pancreatic B-cells up to 12 months after diagnosis. These results are useful when designing trials for therapies aimed to prevent the progression of B-cell destruction in recent-onset T1DM. [source] Demographic and risk factors in patients with head and neck tumorsJOURNAL OF MEDICAL VIROLOGY, Issue 5 2009Ruth Tachezy Abstract The association between human papillomavirus (HPV) infection and the development of head and neck cancer has been documented recently. In this study on 86 head and neck cancer patients and 124 controls, data regarding demographics, behavioral risk factors, and risks related to HPV exposure were collected. HPV detection was carried out using polymerase chain reaction in the tumors and in oral exfoliated cells, and HPV typing by a reverse line blot assay specific for 37 HPV types. Sera were tested by an enzyme-linked immunosorbent assay specific for HPV proteins. Head and neck cancer cases report significantly more oral-anal contact (P,=,0.02) and tobacco and alcohol use than controls (P,=,0.001; P,=,0.02, respectively). High-risk HPV DNA was detected in 43% of oral washings of cases and 4% of controls (P,<,0.0001). The association between the presence of high-risk HPV DNA in oral exfoliated cells and in tumor tissues was statistically significant (adjusted P,<,0.0001). The prevalence of HPV-specific antibodies was significantly higher in cases than in controls (adjusted P,<,0.0001). These results provide epidemiological and immunological evidence for HR HPV as a strong risk factor (OR,=,44.3, P,<,0.0001) for head and neck cancer, even after controlling for age, tobacco and alcohol use. The detection of high-risk HPV DNA in oral exfoliated cells and HPV-specific antibodies in serum can be considered as clinically relevant surrogate markers for the presence of a HPV-associated head and neck cancer, with a high sensitivity (83%) and specificity (88%). J. Med. Virol. 81:878,887, 2009. © 2009 Wiley-Liss, Inc. [source] Re-analysis of 178 previously unidentifiable Mycobacterium isolates in the Netherlands in 1999,2007CLINICAL MICROBIOLOGY AND INFECTION, Issue 9 2010J. Van Ingen Clin Microbiol Infect 2010; 16: 1470,1474 Abstract Nontuberculous mycobacteria (NTM) that cannot be identified to the species level by reverse line blot hybridization assays and sequencing of the 16S rRNA gene comprise a challenge for reference laboratories. However, the number of 16S rRNA gene sequences added to online public databases is growing rapidly, as is the number of Mycobacterium species. Therefore, we re-analysed 178 Mycobacterium isolates with 53 previously unmatched 16S rRNA gene sequences, submitted to our national reference laboratory in 1999,2007. All sequences were again compared with the GenBank database sequences and the isolates were re-identified using two commercially available identification kits, targeting separate genetic loci. Ninety-three out of 178 isolates (52%) with 20 different 16S rRNA gene sequences could be assigned to validly published species. The two reverse line blot assays provided false identifications for three recently described species and 22 discrepancies were recorded in the identification results between the two reverse line blot assays. Identification by reverse line blot assays underestimates the genetic heterogeneity among NTM. This heterogeneity can be clinically relevant because particular sub-groupings of species can cause specific disease types. Therefore, sequence-based identification is preferable, at least at the reference laboratory level, although the exact targets needed for clinically useful results remain to be established. The number of NTM species in the environment is probably so high that unidentifiable clinical isolates should be given a separate species status only if this is clinically meaningful. [source] Distribution of genotypes and antibiotic resistance genes among invasive Streptococcus agalactiae (group B streptococcus) isolates from Australasian patients belonging to different age groupsCLINICAL MICROBIOLOGY AND INFECTION, Issue 3 2008Z. Zhao Abstract Serotype distribution and antibiotic resistance (AR) among group B streptococci (GBS) affect GBS disease prevention strategies, but vary among patient groups. A multiplex PCR-based reverse line blot (mPCR/RLB) hybridisation assay was used to compare the distributions of GBS serotypes, serotype III subtypes and AR-associated genes among 666 invasive isolates from 663 patients, divided into five age groups: infants, early-onset (EO; 0,6 days) and late-onset (LO; 7,90 days); children (aged 3 months to 14 years); women of childbearing age (WCBA; aged 15,45 years); and other adults (males aged >15 years; females aged >45 years). Serotypes Ia and V and serosubtype III-1 accounted for 60% of infections. Serosubtype III-2, which corresponds to a virulent clone belonging to sequence type (ST)17, was relatively uncommon overall (7%), but was associated strongly with LO infant infections, in which it was significantly more common than in adult infections (25/104 (24%) vs. 9/392 (2%), p <0.0001) or in EO infections (25/104 (24%) vs. 14/155 (9%), p <0.005). Erythromycin resistance genes were found in 8% of all isolates (ermB 3%, ermA 2.5% and mefA/E 2%), in 11,15% of isolates of serotypes II and V and subtype III-1, but in none of the isolates of serosubtype III-2 (III-2, 0/49 vs. all others, 54/618 (9%), p <0.04). In summary, the virulent serosubtype III-2 was associated strongly with LO infant GBS infection, but was less likely than other serotypes or serosubtype III-1 to carry AR genes. [source] Use of a multiplex PCR-based reverse line blot (mPCR/RLB) hybridisation assay for the rapid identification of bacterial pathogensCLINICAL MICROBIOLOGY AND INFECTION, Issue 2 2008Y. Wang Abstract The aim of this study was to develop a sensitive and reliable method for the molecular identification of pathogenic bacteria. A multiplex PCR-based reverse line blot (mPCR/RLB) hybridisation assay was developed and evaluated for the rapid identification of 24 systemic and respiratory bacterial pathogens in routine diagnosis. All species-specific probes designed for the RLB hybridised with amplified DNA only from the corresponding species. Sensitivity limits of the mPCR/RLB assay varied among the 24 target organisms from 0.05 pg to 0.5 ng of genomic DNA. The sensitivity of the assay was 2 × 102 CFU/mL for Streptococcus pneumoniae and 6 × 102 CFU/mL for Escherichia coli. The specificity of each probe was tested against 24 species. There were no cross-reactions among any of the 43 probes. The mPCR/RLB assay appeared to be a useful alternative tool for the molecular identification of common pathogens. [source] Identification of novel DNA methylation markers in cervical cancer,INTERNATIONAL JOURNAL OF CANCER, Issue 1 2008Hung-Cheng Lai Abstract Testing for DNA methylation has potential in cancer screening. Most previous studies of DNA methylation in cervical cancer used a candidate gene approach. The aim our study was to identify novel genes that are methylated in cervical cancers and to test their potential in clinical applications. We did a differential methylation hybridization using a CpG island (CGI) microarray containing 8640 CGI tags to uncover methylated genes in squamous cell carcinomas (SCC) of the uterine cervix. Pooled DNA from cancer tissues and normal cervical swabs were used for comparison. Methylation-specific polymerase chain reaction, bisulfite sequencing and reverse transcription polymerase chain reaction were used to confirm the methylation status in cell lines, normal cervices (n = 45), low-grade lesions (n = 45), high-grade lesions (HSIL; n = 58) and invasive squamous cell carcinomas (SCC; n = 22 from swabs and n = 109 from tissues). Human papillomavirus (HPV) was detected using reverse line blots. We reported 6 genes (SOX1, PAX1, LMX1A, NKX6-1, WT1 and ONECUT1) more frequently methylated in SCC tissues (81.5, 94.4, 89.9, 80.4, 77.8 and 20.4%, respectively) than in their normal controls (2.2, 0, 6.7, 11.9, 11.1 and 0%, respectively; p < 0.0001). Parallel testing of HPV and PAX1 methylation in cervical swabs confers an improved sensitivity than HPV testing alone (80% vs. 66%) without compromising specificity (63% vs. 64%) for HSIL/SCC. Testing PAX1 methylation marker alone, the specificity for HSIL/SCC is 99%. The analysis of these novel DNA methylations may be a promising approach for the screening of cervical cancers. © 2008 Wiley-Liss, Inc. [source] | ||||||||||