Usefulness of Mycobacterium tuberculosis molecular typing in a tuberculosis low-endemic agro-industrial setting of Brazil

To highlight the transmission and major phylogenetic clades of Mycobacterium tuberculosis, a retrospective study was carried out at two health facilities in a small agro-industrial area in São Paulo, Brazil, that has a low tuberculosis incidence rate. IS6110-RFLP and spoligotyping were performed on the isolates, with the former revealing that 31.3% (35/112) of strains were clustered. Epidemiological links were found in 16 of the 35 clustered patients and were associated with transmission among patients living in public housing. Spoligotyping grouped 62.8% of the strains. The T genetic family predominated among the isolates. Of interest is that five strains had a pattern characteristic of African or Asian origin (ST535), and two others were of the rare localized type ST1888 (BRA, VEN). In addition, three new types-1889, 1890, and 1891-were identified. Spoligotyping showed that some ST may be circulating to or from Brazil, and RFLP revealed ongoing transmission in inadequately ventilated public-housing buildings. This may point to a failure in tuberculosis control policy.

Molecular differentiation of Salmonella Gallinarum and Salmonella Pullorum by RFLP of fliC gene from Brazilian isolates

Although Salmonella Pullorum and Salmonella Gallinarum cause different diseases in poultry, they are very similar. Both are non-motile and present the same somatic antigenic structure. They are differentiated by biochemical tests. Certain atypical strains are very difficult to distinguish. They do not produce the expected results when dulcitol and ornithine descarxboxylase tests are performed. Therefore, additional tests could be helpful. Many studies have chose the part I of the gene that encodes flagellin (fliC) to differentiate serotypes. Most Salmonella strains have two structural genes (fliC and fliB) that encode flagellins. Non-motile strains generally present these structural genes, but are not able to build a functional flagellum. It was demonstrated that enzymatic restriction of the amplified fliC gene using Hinp1I enzyme can differentiate SG from SP. In the present study, this method was adopted to analyze 14 SP and 22 SG strains, including some strains with atypical results in biochemical tests assessing the utilization of dulcitol and ornithine. The results showed that all SG strains were broken by the enzyme, whereas the 14 SP strains were not.

Restriction fragment length polymorphism (RFLP) in Exon 2 of the BoLA-DRB3 gene in South American cattle

The Bola-DRB3 gene participates in the development of the immune response and is highly polymorphic. For these reasons, it has been a candidate gene in studies of the genetic basis of disease resistance and in population genetic analysis. South American native cattle breeds have been widely replaced by improved exotic breeds leading to a loss of genetic resources. In particular South American native breeds have high levels of fertility and disease resistance. This work describes genetic variability in the BoLA-DRB3 gene in native (Caracu, Pantaneiro, Argentinean Creole) and exotic (Holstein, Jersey, Nelore, Gir) cattle breeds in Brazil and Argentina. PCR-RFLP alleles were identified by combining the restriction patterns for the BoLA-DRB3.2 locus obtained with RsaI, BstY, and HaeIII restriction enzymes. Allelic frequencies and deviations from the Hardy-Weinberg equilibrium were also calculated. Analysis of the 24 BoLA-DRB3 PCR-RFLP alleles identified showed differences in the allele distributions among breeds.

Multidimensional cluster stability analysis from a Brazilian Bradyrhizobium sp RFLP/PCR data set

The taxonomy of the N(2)-fixing bacteria belonging to the genus Bradyrhizobium is still poorly refined, mainly due to conflicting results obtained by the analysis of the phenotypic and genotypic properties. This paper presents an application of a method aiming at the identification of possible new clusters within a Brazilian collection of 119 Bradryrhizobium strains showing phenotypic characteristics of B. japonicum and B. elkanii. The stability was studied as a function of the number of restriction enzymes used in the RFLP-PCR analysis of three ribosomal regions with three restriction enzymes per region. The method proposed here uses Clustering algorithms with distances calculated by average-linkage clustering. Introducing perturbations using sub-sampling techniques makes the stability analysis. The method showed efficacy in the grouping of the species B. japonicum and B. elkanii. Furthermore, two new clusters were clearly defined, indicating possible new species, and sub-clusters within each detected cluster. (C) 2008 Elsevier B.V. All rights reserved.

Typing class I HLA-A gene using a nested PCR-RFLP procedure

In order to detect several new HLA-A class I alleles that have been described since 1998, the original PCR-RFLP method developed to identify the 78 alleles recognized at that time at high resolution level was adapted by us for low and medium resolution levels using a nested PCR-RFLP approach. The results obtained from blood samples of 23 subjects using both the PCR-RFLP method and a commercial kit (MicroSSP1A®, One Lambda Inc.) showed an agreement higher than 95%. The PCR-RFLP adapted method was effective in low and medium resolution histocompatibility evaluations.

Detection of the single nucleotide polymorphism (rs2227307) in the human interleukin 8 gene using a pcr-rflp assay

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

Identification of hybrids between Neotropical fish Leporinus macrocephalus and Leporinus elongatus by PCR-RFLP and multiplex-PCR: Tools for genetic monitoring in aquaculture

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

Genetic identification of the sharks Rhizoprionodon porosus and R. lalandii by PCR-RFLP and nucleotide sequence analyses of 5S rDNA

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

Identification of guitarfish species Rhinobatos percellens, R. horkelli, and Zapteryx brevirostris (Chondrichthyes) using mitochondrial genes and RFLP technique

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

Identification of the shark species Rhizoprionodon lalandii and R-porosus (Elasmobranchii, Carcharhinidae) by multiplex PCR and PCR-RFLP techniques

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

Genetic grouping of avian infectious bronchitis virus isolated in Brazil based on RT-PCR/RFLP analysis of the S1 gene

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

RFLP analysis of genetic variation in species of section Arachis, genus Arachis (Leguminosae)

Four A-genome species of the genus Arachis ( A. cardenasii, A. correntina, A. duranensis, A. kempff-mercadoi), three B genomes species ( A. batizocoi, A. ipaensis and A. magna), the AABB allotetraploid A. hypogaea (cultivated peanut) and introgression lines resulting from a cross between A. hypogaea and A. cardenasii were analyzed by RFLP. The A genome species (cytologically characterized by the presence of a small chromosome pair 'A') were closely similar to each other and shared a large number of restriction fragments. In contrast, the B genome species differed more from one another and shared few fragments. The results of this study indicate that the absence of the small chromosome pair is not a good criterion for grouping species of section Arachis as B genome species, since their genome might be quite distinct from the B genome of A. hypogaea. The lowest genetic variation was detected within accessions of A. duranensis (17 accessions), followed by A. batizocoi (4 accessions) and A. cardenasii (9 plants of accession GKP 10017). The high level of genetic variation found in A. cardenasii might indicate that not all accessions of wild species of Arachis are autogamous, as reported for A. hypogaea.

A novel PCR-RFLP assay for the detection of the single nucleotide polymorphism at position+1440 in the human CXCR2 gene

We designed a novel PCR-RFLP assay to genotype for the CXCR2 +1440 (G/A) single nucleotide polymorphism, which provides a simple, low-cost, practical, and reproducible method. Allele frequencies in healthy Brazilian individuals were found to be 0.65% for allele A and 0.35% for allele G.

Use of PCR-RFLP (Polymerase Chain Reaction-Restricted Fragment Length Polymorphism) in the gene of the enzyme Stearoyl-CoA-Desaturase in Bubalus bubalis

The milk is an important food because it contents Conjugated Linoleic Acids (CIA). These fatty acids are synthesized in mammary gland under action of the enzyme Stearoyl CoA-Desaturase (SCD) and have showed some positive effects in human disease prevention and treatments. A variation of CLA in milk fat exists and can be partially explained by the different levels of expression of SCD. The aim was to study part of the encoding regions of SCD's gene using PCR-RFLP (Polymerase Chain Reaction-Restriction Fragment Length Polymorphism). Genomic DNA was extracted from lactating Murrah females. After this, PCR reactions were made by using primers Z (sic) (sic) D1 that encloses exon I, II and intron I. The fragments amplified are composed by 938 pb. Then, RFLP techniques were applied in the fragments using the restriction enzymes Pst I and Sma I. The enzyme Pst I has generated fragments of 788pb and 150bp and the Sma I has generated fragments of 693pb and 245pb. All the animals showed the same migration standard for both enzymes, characterizing a genetic monomorphism for this region of SCD gene. The analysis determined that there aren't genetic differences between these animals in the studied regions by using Pst I and Sma I enzymes.