Genotyping of hepatitis C virus performed by type-specific PCR in comparison to nucleotide sequencing of NS5 and core regions.
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Genotype↗
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CTLA4/CT60 polymorphism is not relevant in susceptibility to autoimmune inflammatory intestinal disorders.
The aim of this work was to investigate the possible influence of the recently described CT60 A/G dimorphism of the CTLA4 (cytotoxic T-lymphocyte antigen 4) gene in the susceptibility to two different autoimmune inflammatory intestinal disorders, inflammatory bowel disease (IBD) and celiac disease. We analyzed a case-control cohort composed of 528 Spanish patients with IBD (284 with Crohn disease and 244 with ulcerative colitis) and 454 unrelated healthy individuals, and additionally a group of 90 celiac disease families. CT60 genotyping was performed with a TaqMan 5' allelic discrimination assay. After comparing patients with IBD with the control population, we found no significant deviation in the distribution of the alleles or genotypes of CTLA4/CT60 dimorphism. In addition, by means of familial and case-control analysis, no evidence for a statistically significant association was observed between CTLA4/CT60 and celiac disease susceptibility. Therefore, our results suggest that the CTLA4/CT60 polymorphism does not play a major role in inflammatory intestinal disorders.
Antigens, CD↗
Chromosome 5 and Parkinson disease.
Parkinson disease (PD) is the second most common neurodegenerative disorder. Despite the identification of five causative genes, the majority of PD etiology is still unknown. A region on chromosome 5q is one of the few regions of the genome found linked in multiple studies of familial PD. Analyses were performed using genotypic data from two independent research studies to evaluate rigorously the evidence of linkage on chromosome 5. The combined sample consisting of 1238 affected individuals from 569 multiplex PD families were genotyped for a common set of 20 microsatellite markers spanning an 80 cM region on chromosome 5q. Two disease models were employed and model-free linkage analyses were performed to detect linkage to a PD susceptibility gene and also to detect linkage to a quantitative phenotype, age of onset of PD. There was little evidence of linkage using either a narrower or broader disease definition (lod <0.5). Analyses employing age of onset of PD as the phenotype produced a lod score of 1.8. These results in a very large sample of familial PD suggest that it is unlikely that a PD susceptibility gene is located on chromosome 5q. Evidence for a locus contributing to the age of onset of PD is modest at best (empirical P-value=0.07).
Adult↗
Characterization of Chlamydia trachomatis omp1 genotypes among sexually transmitted disease patients in Sweden.
A method for detection and genotyping of genital Chlamydia trachomatis infections based on omp1 gene amplification and sequencing was developed. DNA was extracted from urogenital or urine samples using a Chelex-based method, and an approximately 1,100-bp-long fragment from the omp1 gene was directly amplified and sequenced. Genotyping was performed by BLAST similarity search, and phylogenetic tree analysis was used to illustrate the evolutionary relationships between clinical isolates and reference strains. The method was used to determine the genotypes of C. trachomatis in 237 positive urogenital and/or urine specimens collected at a Swedish sexually transmitted disease clinic during 1 year. The most common genotypes corresponded to serotypes E (47%) and F (17%). The omp1 gene was highly conserved for genotype E (106 of 112 samples without any mutation) and F (41 of 42 samples without any mutation) strains but appear slightly less conserved for genotypes G (n = 6) and H (n = 6), where the sequences displayed one to four nucleotide substitutions relative to the reference sequence. Genotyping of samples collected at the follow-up visit indicated that two patients had become reinfected, while three other patients suffered treatment failure or reinfection. One woman appeared to have a mixed infection with two different C. trachomatis strains. This omp1 genotyping method had a high reproducibility and could be used for epidemiological characterization of sexually transmitted Chlamydia infections.
Adolescent↗
[Maintenance dose requirement for phenytoin is lowered in genetically impaired drug metabolism independent of concommitant use of other antiepileptics].
OBJECTIVE: To investigate the effect of genetically determined impaired drug metabolism and of the use of comedication on phenytoin maintenance dose requirement. DESIGN: Descriptive. METHOD: In 60 patients on long-term phenytoin therapy the concentration of phenytoin in serum was measured and CYP2C9 genotyping was performed (mutant alleles of CYP2C9 are associated with impaired phenytoin metabolism). In addition, the use of other antiepileptics concurrently with phenytoin was reviewed: phenobarbital, carbamazepine and valproic acid. CYP2C9 genotype and comedication were connected to phenytoin daily dose requirement. The 60 patients were 38 men and 22 women, between 16 and 74 years of age, and all mentally disabled. RESULTS: Genotyping revealed that of the 60 patients 38% (n = 23) carried at least one mutant CYP2C9 allele. Their mean dose of phenytoin was 199 mg dd, while the mean required maintenance dose in non-carriers (n = 37) was 287 mg dd (p < 0.01). When the use of comedication was taken into account, it appeared that the mean phenytoin daily doses in the different comedication groups did not differ significantly. CONCLUSION: Patients with a genetically determined impaired phenytoin metabolism required on average a 30% lower dose than those with a normal metabolism, while the concomitant use of other antiepileptics appeared to have no effect on phenytoin dose requirement.
Adolescent↗
Tumor necrosis factor-alpha gene polymorphism in severe and mild-moderate rheumatoid arthritis.
OBJECTIVE: To examine whether severe rheumatoid arthritis (RA) carries a -238 or +489 tumor necrosis factor-alpha (TNF-alpha) genotype different from mild-moderate RA. METHODS: We investigated 163 patients (66 with severe disease) and 67 healthy blood donor controls. Genotyping was performed by polymerase chain reaction-restriction fragment length polymorphism. RESULTS: Patients with severe RA (all active disease despite disease modifying antirheumatic drug combination therapy) disclosed the -238 GG genotype in 100% of cases versus 92.8% of the mild-moderates and 92.5% of controls (OR 11.7, Cl 0.6-216, p = 0.03). The +489 AA genotype was seen less often in patients than in controls (OR 4.2. CI 0.97-18.4, p = 0.045), and the contribution to this trend appeared predominant in the anti-TNF treated subgroup. CONCLUSION: The -238 AG genotype was absent in severe RA; in contrast, patients with mild-moderate RA disclosed the same frequency as controls. Thus -238 GG homozygosity is associated with severe RA. The +489 AA genotype might instead protect against worse outcome in RA.
Adolescent↗
First trimester prenatal diagnosis of 21-hydroxylase deficiency by linkage analysis to HLA-DNA probes and by 17-hydroxyprogesterone determination.
The close genetic linkage between the gene for congenital adrenal hyperplasia due to 21-hydroxylase (21-OH) deficiency and HLA genes allowed us to use the polymorphism of this system as a marker of the disease. HLA genotyping can be performed by using restriction enzyme fragments hybridized with specific probes instead of serologic methods. In seven pregnancies at risk for 21-OH deficiency, a first trimester prenatal diagnosis has been performed by determining the fetal genotype by linkage analysis of DNA from chorionic villi using HLA class I and class II probes. In four of these pregnancies, determination of 17-OH progesterone in first trimester amniotic fluid afforded a complementary approach to the diagnosis.
17-alpha-Hydroxyprogesterone↗
Prevalence of the Factor V G1691A and the Factor II/prothrombin G20210A gene polymorphisms among Tamilians.
We have investigated the prevalence of the Factor II G20210A and Factor V G1691A single nucleotide polymorphisms (SNPs) in a South Indian-Tamil Nadu population. The SNP genotyping was performed using a polymerase chain reaction (PCR)/restriction fragment length polymorphism analysis and by a recently FDA-approved LightCycler real-time PCR assay. Of 72 samples that were genotyped, 4 (5.5%) patients were heterozygous for the Factor V SNP and no homozygous mutant patients were identified. None of the patients were shown to be either heterozygous or homozygous mutant for the Factor II SNP. All samples showed 100% concordance between the PCR/RFLP assay and the LightCycler assay. While this study identified the prevalence of the Factor V SNP to be similar to that of other reported populations, the absence of the Factor II allele is consistent with reports in more isolated populations. In addition, the results of this study do not support a role for these SNPs in acute myocardial infarction in the Tamilian population.
DNA Mutational Analysis↗
Analysis of two common alpha 1-antitrypsin deficiency alleles (PI*Z and PI*S) in subjects with periodontitis.
BACKGROUND: pi alpha 1-Antitrypsin deficiency is a genetically determined condition resulting in predisposition to certain inflammatory diseases due to a protease: antiprotease imbalance that is exacerbated by tobacco smoking. Limited evidence suggests that there may be a significant enrichment of mild alpha 1-antitrypsin deficiency phenotypes in subjects with chronic inflammatory periodontal disease. OBJECTIVE: To examine the prevalence of two common alpha1-antitrypsin deficiency alleles (PI*Z and PI*S) in a UK population of subjects with periodontitis. SUBJECTS AND METHODS: The prevalence of PI*M, PI*S and PI*Z allele combinations was determined in 31 subjects with periodontitis and compared with 31 healthy control subjects matched for smoking status, ethnicity, age and gender. alpha 1-Antitrypsin genotyping was performed by multiplex real-time fluorescence polymerase chain reaction (PCR) using DNA extracted from whole blood. RESULTS: There was no difference in the proportion of any alpha 1-antitrypsin genotype found in the diseased and control populations. CONCLUSIONS: We did not find evidence to support an association between mutant PI* alleles and periodontitis in a small, controlled study. Larger studies will be required to clarify the relationship between alpha1-antitrypsin genotype and susceptibility to inflammatory periodontal disease.
Adult↗
Polymorphisms in the TNF-alpha and TNF-receptor genes in patients with coronary artery disease.
BACKGROUND: Coronary artery disease (CAD) is both multifactorial and polygenic in nature. Atheroma formation, the pathological hallmark of CAD, is an inflammatory process, with pro-inflammatory cytokines, such as tumour necrosis factor-alpha (TNF-alpha), having a major role in its pathogenesis. We have therefore investigated whether polymorphisms in the TNF-alpha (- 238 and - 308), TNF receptor 1 (position - 609 and + 10, intron 6) and TNF receptor 2 (position + 422, codon 198) genes show an association with CAD. MATERIALS AND METHODS: Patients with angiographically proven single vessel (n = 58) and multivessel (n = 122) CAD were compared to patients with angiographically proven normal coronary arteries (n = 79) and volunteers without clinical evidence of CAD (n = 250). Genotyping was performed by PCR-RFLP analysis. For the TNF-alpha polymorphisms, a meta-analysis of all published studies was also undertaken. RESULTS: No significant differences in allele or genotype frequencies were found between the normal coronary artery group or healthy volunteers and patients with CAD for any of the polymorphisms. There was also no difference in allele frequency between patients with single- and multivessel disease. For the - 308 and - 238 TNRalpha gene polymorphisms, a meta-analysis of our data and previously published studies failed to demonstrate any significant association with CAD. CONCLUSIONS: Polymorphisms in the TNF-alpha promoter region and TNF-receptor genes are not associated with the development of CAD.
Adult↗
Clinical utility of HIV-1 genotyping and expert advice: the Havana trial.
OBJECTIVE: To determine whether HIV-1 genotyping and expert advice add additional short-term virologic benefit in guiding antiretroviral changes in HIV+ drug-experienced patients. DESIGN: A two factorial (genotyping and expert advice), randomized, open label, multi-center trial. The patients were stratified according to the number of treatment failures. PATIENTS AND METHODS: HIV-1 infected patients on stable antiretroviral therapy who presented virological failure were included into the study. Genotypic testing was performed by using TrueGene HIV Genotyping kit and the results were interpreted by a software package (RetroGram, version 1.0). An expert advisory committee suggested the new therapeutic approach based on clinical information alone or on clinical information plus HIV-1 genotyping results. Plasma HIV-1 RNA load, CD4+ cell count and adverse events were recorded at baseline and every 12 weeks. RESULTS: A total of 326 patients were included. The baseline CD4+ cell count and plasma HIV-1 RNA were 387 (+/- 224) x 10(6) cells/l and 4 (+/- 1) log(10) respectively. The proportion of patients with plasma HIV-1 RNA < 400 copies/ml at 24 weeks differed between genotyping and no genotyping arms (48.5 and 36.2%, P < 0.05). Factors associated with a higher probability of plasma HIV-1 RNA < 400 copies/ml were HIV-1 genotyping [odds ratio (OR), 1.7; 95% confidence interval (CI), 1.1-2.8; P = 0.016] and the expert advice in patients failing to a second-line antiretroviral therapy (OR, 3.2; 95% CI, 1.2-8.3; P = 0.016). CONCLUSIONS: HIV-1 genotyping interpreted by a software package improves the virological outcome when it is added to the clinical information as a basis for decisions on changing antiretroviral therapy. The expert advice also showed virologic benefit in the second failure group.
Adult↗
Similar genotypes of Pneumocystis jirovecii in different forms of Pneumocystis infection.
This study describes the genotyping of Pneumocystis jirovecii organisms isolated from three groups of patients that developed diverse forms of P. jirovecii infection; the patients were monitored in the same French hospital. Forty archival specimens from 13 adults with Pneumocystis pneumonia, eight adults colonized by P. jirovecii and 19 immunocompetent infants infected with the fungus contemporaneously with a bronchiolitis episode were analysed retrospectively. Genotyping was performed by analysis of sequences of the internal transcribed spacer (ITS)1 and ITS2 regions, and of the dihydropteroate synthase (DHPS) locus. At the ITS regions, a high diversity of genotypes, identical main genotypes (B(1)a(3) and B(2)a(1)) and the occurrence of mixed infections (more than one genotype) were observed in the three patient groups. At the DHPS locus, the results indicated the presence of mutants in the two adult groups, as well as in the infant group. Consequently, at these two independent genomic regions, P. jirovecii isolates from patients who developed different forms of infection and who lived in the same geographical region presented common characteristics. These results suggest that patients infected with P. jirovecii, whatever the form of infection they present, are part of a common human reservoir for P. jirovecii.
Adult↗
Optimal diversity-dependent contributions of genotypes to mixtures.
Deployment of genotypes to a production population decisively depends on the measure of diversity, a consideration that parallels genetic gain in the management of genotype mixtures. Optimal diversity-dependent deployment has been developed in this study for a family of diversity indices in the genetical and ecological context. The optimal solution at given diversity was expressed as the relationship between genotype contributions and their genetic performances, which maximized genetic gain. Numerical calculations were performed by using genotypes generated from normal order statistics. An optimal deployment in one situation could be nonoptimum in another. Classical uniform deployment, where superior genotypes equally contribute to the mixtures, was the limit of optimal deployment. Comparisons were made between optimal and uniform deployment and between optimal and nonoptimal deployment where genotypes contributed proportionally to the mixtures in accordance to their genetic superiority. The superiority in gain of optimal deployment over that of uniform deployment increased as the difference between the diversity measure under optimal deployment and the contributing number (N) of genotypes under uniform deployment became large and as the diversity measure and N under optimal deployment increased. The superiority over nonoptimal deployment increased rapidly at low diversity, reaching a maximum somewhere at diversity between 1 and N. Scale of superiority depended on the similitude between optimum and nonoptimum deployment; the larger the distinctiveness, the greater the superiority.
Agriculture↗
Clinical features of sarcoidosis in relation to HLA distribution and HLA-DRB3 genotyping by PCR-RFLP.
BACKGROUND: Susceptibility to the development of sarcoidosis has been demonstrated to be associated with HLA-DR5, -DR6, and -DR8 encoded by the DRB1 gene. However, involvement of the DRB3 (HLA-DR52) gene in the development of sarcoidosis remains unclear. METHODS: HLA-DRB3 genotyping was performed using the PCR-RFLP method and the clinical features of the patients with and without the DR3, 5, 6, 8 group antigens were compared. RESULTS: HLA-DRB3 genotyping indicated an association between DRB3*0101 and sarcoidosis. The DR8 haplotype lacking the DRB3 gene has been found to be increased significantly in sarcoidosis, suggesting that the HLA-DRB3 gene is not a primary determinant of predisposition to sarcoidosis. The association of DRB3*0101 with sarcoidosis is attributable to linkage disequilibrium with DR5- and DR6-associated alleles. There were significant decreases in the DR3, 5, 6, 8 group (DR5, DR6, or DR8) antigen frequencies in patients with retinal perivasculitis, high intraocular pressure (or secondary glaucoma), and optic nerve and/or macular lesion. Correlations were observed among the DR3, 5, 6, 8 group antigens, early onset sarcoidosis and disease with fewer intraocular lesions. CONCLUSION: This established a molecular basis for some of the clinical heterogeneity observed in sarcoidosis.
Adolescent↗
The association of single nucleotide polymorphisms in the 5'-regulatory region of the lumican gene with susceptibility to high myopia in Taiwan.
PURPOSE: To study the relationships between single nucleotide polymorphisms (SNPs) of lumican, decorin, and DSPG3 genes and high myopia. METHODS: One hundred and twenty adult patients with high myopia (< -10.0 D) and 137 controls were used to study the relationships between the decorin, lumican, and DSPG genes and high myopia. All subjects were free of ocular diseases, other than myopia, as well as of other systemic genetic diseases. Genotyping was performed by direct sequencing after PCR amplification of chromosomal DNA. Allele frequencies were tested for Hardy-Weinberg disequilibrium. The chi(2) or Fisher test was conducted to investigate the genotypic and allelic distribution between the high myopia and control groups. RESULTS: The genotyping success rate was 100%. Univariate analysis revealed significant differences between patients and control subjects with respect to one of the SNPs (rs3759223, C->T) of the lumican gene, with a p value of 0.000283. There was no significant relationship between other SNPs of lumican, decorin, and DSPG genes and high myopia. CONCLUSIONS: Our results indicate that an SNP (rs3759223), which is located in the promoter region of the lumican gene, may be worth further investigation to determine its association with development of high myopia.
5' Flanking Region↗
Glutathione-S-transferase genotypes and the adverse effects of azathioprine in young patients with inflammatory bowel disease.
BACKGROUND: Adverse drug reactions to azathioprine, the prodrug of 6-mercaptopurine, occur in 15%-38% of patients and the majority are not explained by thiopurine-S-methyltransferase (TPMT) deficiency. Azathioprine is known to induce glutathione depletion and consumption of glutathione is greater in cells with high glutathione-S-transferase (GST) activity compared with those with low activity; moreover, some reports indicate that GST might play a direct role in the reaction of glutathione with azathioprine. The association between polymorphisms of GST-M1, GST-P1, GST-T1, and TPMT genes and the adverse effects of azathioprine was therefore investigated. METHODS: Seventy patients with inflammatory bowel disease (IBD), treated with azathioprine, were enrolled and clinical data were retrospectively determined. TPMT and GST genotyping were performed by polymerase chain reaction (PCR) assays on DNA extracted from blood samples. RESULTS: Fifteen patients developed adverse effects (21.4%); there was a significant underrepresentation of the GST-M1 null genotype among patients developing adverse drug reactions to azathioprine (odds ratio [OR] = 0.18, 95% confidence interval [CI] = 0.037-0.72, P = 0.0072) compared with patients who did not develop adverse effects. Patients heterozygous for TPMT mutations presented a marginally significant increased probability of developing adverse effects (OR = 6.38, 95% CI = 0.66-84.1, P = 0.062). Moreover, among the 55 patients who did not develop adverse effects, there was a significant underrepresentation of the GST-M1 null genotype among patients who displayed lymphopenia as compared with those that did not display this effect of azathioprine (OR = 0.15, 95% CI = 0.013-1.08, P = 0.032). CONCLUSION: Patients with IBD with a wildtype GST-M1 genotype present increased probability of developing adverse effects and increased incidence of lymphopenia during azathioprine treatment.
Adolescent↗
No association between the C-1562T polymorphism in the promoter of matrix metalloproteinase-9 gene and non-small cell lung carcinoma.
The matrix metalloproteinases (MMPs) are a family of highly conserved metal-dependent proteolytic enzymes, their main function is to degrade different components of extracellular matrix (ECM). Moreover, they play roles in regulation of cell growth, apoptosis, angiogenesis and immune surveillance. Natural sequence variations in the MMP genes may result in differential expression of MMPs in different individuals and therefore may be associated with the development and progression of diseases. The aim of this study is to assess the effects of the C-1562T polymorphism in the MMP-9 promoter on the risk of occurrence and lymphatic metastasis of non-small cell lung carcinoma (NSCLC). The MMP-9 genotyping was performed in 243 pathologically diagnosed NSCLC patients and 350 healthy controls without overt cancer by using polymerase chain reaction-restriction fragment length polymorphism analysis. The distribution of the MMP-9 genotypes in NSCLC patients and healthy controls was in consistent with Hardy-Weinberg equilibrium. The frequency of the C/C, C/T and T/T genotypes in healthy controls was 79.4, 20.6 and 0%, respectively. Neither the overall genotype nor allelotype distribution in NSCLC patients showed significant difference from that in healthy controls (P=0.21 and 0.43, respectively). Compared with the C/C genotype, genotypes with the T allele did not show significant influence on the risk of NSCLC development (age and gender adjusted OR=1.13, 95% CI=0.76-1.68). Stratification by onset age, smoking status and tumor histological type also showed no association between the MMP-9 polymorphism and the risk of NSCLC. Furthermore, the genotype distribution between NSCLC patients with and without lymphatic metastasis was not significantly different. Therefore, the present study suggests that the MMP-9 C-1562T polymorphism may not be used as a useful marker to predicate susceptibility and lymphatic metastasis in NSCLC.
Adenocarcinoma↗
Platelet serotonin transporters and the transporter gene in control subjects, unipolar patients and bipolar patients.
OBJECTIVE: The purpose of the present study was to relate the number of platelet serotonin transporters in unipolar and bipolar patients and in control subjects to two polymorphisms in the serotonin transporter gene: a VNTR in intron 2 and a deletion/insertion in the promoter region. METHOD: Density of platelet serotonin transporters was determined by radioligand binding analysis. Genotyping was performed by PCR amplification of polymorphic regions followed by size determination of the obtained fragments. RESULTS: The control subjects and the two groups of patients were similar with respect to the genotype and allele distribution belonging to the two polymorphisms in the serotonin transporter gene for. An interaction between status (control, unipolar- or bipolar patient) and VNTR genotype regarding the number of platelet serotonin transporters was observed; unipolar patients with the genotype 12/10 had more platelet serotonin transporters than bipolar patients and controls with this genotype. No association related to the polymorphism was found in the promoter region of the serotonin transporter gene. CONCLUSION: An association was observed between the polymorphism in intron 2 of the serotonin transporter gene and the number of platelet serotonin transporters. Unipolar patients with a particular genotype had more platelet serotonin transporters than the corresponding controls and bipolar patients.
Adult↗