Association of ABO gene mutations resulting in a rare B subgroup

Background and Objectives B subgroups are rare and the genetic analysis reported to date has been limited.Materials and Methods Serological and molecular investigations were performed in blood from a B-subgroup donor.Results Red cells did not react with anti-B and anti-AB reagents. However, cells absorbed anti-B. Red cells presented positive reactions with anti-H, and saliva secreted H substance. The molecular study demonstrated a B allele with the substitutions 467C>T, 646T>A, 681G>A, 771C>T, 796C>A, 803G>C, 829G>A and an 0 allele with the sequence of 002.Conclusions It is probable that the presence in exon 7 of some of the 002 substitutions could have weakened the enzymatic activity of the encoded B transferase.

Plasmid-based controls to detect rpoB mutations in Mycobacterium tuberculosis by quantitative polymerase chain reaction-high-resolution melting

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

No mutations found in exons of TP53, H-RAS and K-RAS genes in liver of male Wistar rats submitted to a medium-term chemical carcinogenesis assay

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Variability and resistance mutations in the hepatitis C virus NS3 protease in patients not treated with protease inhibitors

The goal of treatment of chronic hepatitis C is to achieve a sustained virological response, which is defined as exhibiting undetectable hepatitis C virus (HCV) RNA levels in serum following therapy for at least six months. However, the current treatment is only effective in 50% of patients infected with HCV genotype 1, the most prevalent genotype in Brazil. Inhibitors of the serine protease non-structural protein 3 (NS3) have therefore been developed to improve the responses of HCV-infected patients. However, the emergence of drug-resistant variants has been the major obstacle to therapeutic success. The goal of this study was to evaluate the presence of resistance mutations and genetic polymorphisms in the NS3 genomic region of HCV from 37 patients infected with HCV genotype 1 had not been treated with protease inhibitors. Plasma viral RNA was used to amplify and sequence the HCV NS3 gene. The results indicate that the catalytic triad is conserved. A large number of substitutions were observed in codons 153, 40 and 91; the resistant variants T54A, T54S, V55A, R155K and A156T were also detected. This study shows that resistance mutations and genetic polymorphisms are present in the NS3 region of HCV in patients who have not been treated with protease inhibitors, data that are important in determining the efficiency of this new class of drugs in Brazil.

No mutations found in exon 2 of gene p16CDKN2A during rat tongue carcinogenesis induced by 4-nitroquinoline-1-oxide

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Mutations in IRF6 cause Van der Woude and popliteal pterygium syndromes

Interferon regulatory factor 6 (IRF6) belongs to a family of nine transcription factors that share a highly conserved helix-turn-helix DNA-binding domain and a less conserved protein-binding domain. Most IRFs regulate the expression of interferon-alpha and -beta after viral infection(1), but the function of IRF6 is unknown. The gene encoding IRF6 is located in the critical region for the Van der Woude syndrome (VWS; OMIM 119300) locus at chromosome 1q32-q41 (refs 2,3). The disorder is an autosomal dominant form of cleft lip and palate with lip pits(4), and is the most common syndromic form of cleft lip or palate. Popliteal pterygium syndrome (PPS; OMIM 119500) is a disorder with a similar orofacial phenotype that also includes skin and genital anomalies(5). Phenotypic overlap(6) and linkage data(7) suggest that these two disorders are allelic. We found a nonsense mutation in IRF6 in the affected twin of a pair of monozygotic twins who were discordant for VWS. Subsequently, we identified mutations in IRF6 in 45 additional unrelated families affected with VWS and distinct mutations in 13 families affected with PPS. Expression analyses showed high levels of Irf6 mRNA along the medial edge of the fusing palate, tooth buds, hair follicles, genitalia and skin. Our observations demonstrate that haploinsufficiency of IRF6 disrupts orofacial development and are consistent with dominant-negative mutations disturbing development of the skin and genitalia.

Prevalence and nonrandom distribution of exonic mutations in interferon regulatory factor 6 in 307 families with Van der Woude syndrome and 37 families with popliteal pterygium syndrome

Purpose: Interferon regulatory factor 6 encodes a member of the IRF family of transcription factors. Mutations in interferon regulatory factor 6 cause Van der Woude and popliteal pterygium syndrome, two related orofacial clefting disorders. Here, we compared and contrasted the frequency and distribution of exonic Mutations in interferon regulatory factor 6 between two large geographically distinct collections of families with Van der Woude and between one collection of families with popliteal pterygium syndrome. Methods: We performed direct sequence analysis of interferon regulatory factor 6 exons oil samples from three collections, two with Van der Woude and one with popliteal pterygium syndrome. Results: We identified mutations in interferon regulatory factor 6 exons in 68% of families in both Van der Woude collections and in 97% of families with popliteal pterygium syndrome. In sum, 106 novel disease-causing variants were found. The distribution of mutations in the interferon regulatory factor 6 exons in each collection was not random; exons 3, 4, 7, and 9 accounted for 80%. In the Van der Woude collections, the mutations were evenly divided between protein truncation and missense, whereas most mutations identified in the popliteal pterygium syndrome collection were missense. Further, the missense mutations associated with popliteal pterygium syndrome were localized significantly to exon 4, at residues that are predicted to bind directly to DNA. Conclusion: The nonrandom distribution of mutations in the interferon regulatory factor 6 exons suggests a two-tier approach for efficient mutation screens for interferon regulatory factor 6. The type and distribution of mutations are consistent with the hypothesis that Van der Woude is caused by haploinsufficiency of interferon regulatory factor 6. Oil the other hand, the distribution of popliteal pterygium syndrome-associated mutations suggests a different, though not mutually exclusive, effect oil interferon regulatory factor 6 function. Genet Med 2009:11(4):241-247.

Gene Mutations in Esophageal Mucosa of Chagas Disease Patients

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Gene Mutations and Polymorphisms of TP53 and FHIT in Chronic Esophagitis and Esophageal Carcinoma

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Antiretroviral resistance mutations in human immunodeficiency virus type 1 infected patients enrolled in genotype testing at the Central Public Health Laboratory, São Paulo, Brazil: preliminary results

Antiretroviral resistance mutations (ARM) are one of the major obstacles for pharmacological human immunodeficiency virus (HIV) suppression. Plasma HIV-1 RNA from 306 patients on antiretroviral therapy with virological failure was analyzed, most of them (60%) exposed to three or more regimens, and 28% of them have started therapy before 1997. The most common regimens in use at the time of genotype testing were AZT/3TC/nelfinavir, 3TC/D4T/nelfinavir and AZT/3TC/efavirenz. The majority of ARM occurred at protease (PR) gene at residue L90 (41%) and V82 (25%); at reverse transcriptase (RT) gene, mutations at residue M184 (V/I) were observed in 64%. One or more thymidine analogue mutations were detected in 73%. The number of ARM at PR gene increased from a mean of four mutations per patient who showed virological failure at the first ARV regimens to six mutations per patient exposed to six or more regimens; similar trend in RT was also observed. No differences in ARM at principal codon to the three drug classes for HIV-1 clades B or F were observed, but some polymorphisms in secondary codons showed significant differences. Strategies to improve the cost effectiveness of drug therapy and to optimize the sequencing and the rescue therapy are the major health priorities.

Identification and characterization of novel mutations of the aspartoacylase gene in non-Jewish patients with Canavan disease

Canavan disease, an inherited leukodystrophy, is caused by mutations in the aspartoacylase (ASPA) gene. It is most common among children of Ashkenazi Jewish descent but has been diagnosed in many diverse ethnic groups. Two mutations comprise the majority of mutant alleles in Jewish patients, while mutations in the ASPA gene among non-Jewish patients are different and more diverse. In the present study, the ASPA gene was analysed in 22 unrelated non-Jewish patients with Canavan disease, and 24 different mutations were found. of these,14 are novel, including five missense mutations (E24G, D68A, D249V, C152W, H244R), two nonsense mutations (Q184X, E214X), three deletions (923delT, 33del13, 244delA), one insertion mutation (698insC), two sequence variations in one allele ([10T>G; 11insG]), an elimination of the stop codon (941A>G, TAG-->TGG, X314W), and one splice acceptor site mutation (IVS1 - 2A>T). The E24G mutation resulted in substitution of an invariable amino acid residue (Glu) in the first esterase catalytic domain consensus sequence. The IVS1 - 2A>T mutation caused the retention of 40 nucleotides of intron 1 upstream of exon 2. The results of transient expression of the mutant ASPA cDNA containing these mutations in COS-7 cells and assays for ASPA activity of patient fibroblasts indicated that these mutations were responsible for the enzyme deficiency. In addition, patients with the novel D249V mutation manifested clinically at birth and died early. Also, patients with certain other novel mutations, including C152W, E214X, X314W, and frameshift mutations in both alleles, developed clinical manifestations at an earlier age than in classical Canavan disease.

pncA mutations in pyrazinamide-resistant Mycobacterium tuberculosis clinical isolates from the southeast region of Brazil

Objectives: To investigate the presence of mutations in the pncA gene in 31 pyrazinamide-resistant Mycobacterium tuberculosis and 5 susceptible strains. MICs and pyrazinamidase (PZase) activity were also determined.Methods: All 36 M. tuberculosis clinical isolates were genotyped by mycobacterial interspersed repetitive units (MIRUs) and most were also typed by spoligotyping. The MIC value necessary to inhibit 99% of the resistant mycobacterial isolates was determined by microplate Alamar Blue assay (MABA) and by Lowenstein-Jensen assay (LJA). The PZase activity was measured by pyrazinamide deamination to pyrazinoic acid and ammonia, and the entire pncA sequence including the 410 by upstream from the start codon was determined by DNA sequencing of purified PCR products.Results: of the 31 isolates resistant to pyrazinamide, 26 (83.9%) showed at least one mutation in the pncA gene or in its putative regulatory region: Among the 22 different mutations detected in the pncA gene and in its regulatory region, 9 (40.9%) mutations (consisting of six substitutions, two insertions and one deletion) have not been described in previous studies. Three pyrazinamide-resistant isolates, confirmed by MIC varying from 800 to 1600 mg/L, carried the wild-type pncA sequence and retained PZase activity.Conclusions: These results contribute to the knowledge of the molecular mechanism of pyrazinamide resistance in Brazil and also expand the profile of pncA mutations worldwide. The MABA was successfully used to determine the MICs of pyrazinamide.