Rodrigo Rodenbusch

Allele frequencies of 15 STRs in a representative sample of the Brazilian population

Allele frequencies for 15 short tandem repeat (STR) loci were obtained from a sample of 12,030 individuals undergoing paternity testing. This sample includes individuals from all States in Brazil, combined according to the current country division into five regions (North, Northeast, Central West, Southeast, and South). The most polymorphic loci were D2S1338 and D18S51. All the analysed loci meet Hardy-Weinberg equilibrium expectations. Combined power of discrimination and combined power of exclusion for the 15 tested STR loci were 0.999999999999990 and 0.9999992, respectively. Comparative analysis between populations from different Brazilian macroregions as well as between Brazil and other relevant populations are presented.

Mutation rate estimates for 13 STR loci in a large population from Rio Grande do Sul, Southern Brazil

Short tandem repeat (STR) polymorphisms have been extensively used in forensic genetics analysis. Knowledge about the locus-specific mutation rates of STRs improves forensic probability calculations and interpretations of diversity data. To incorporate single-locus diversity information into autosomal STR mutation rate estimations, 13 STR loci were studied during 2007–2009 in 10,959 paternity investigation cases from Rio Grande do Sul, the southernmost state of Brazil, covering an overall number of 284,934 allelic transfers. A total of 355 mutations were identified; 348 repeats were gains or losses of one step, three were gains or losses of two steps, and four were gains or losses of not stepwise mutation. The mutation rates ranged from 4.6 × 10−5 to 2.3 × 10−3, and the overall mutation rate estimate was 1.2 × 10−3. The average of the paternal mutation rate (1.8 × 10−3) was five times higher than the maternal rate (0.36 × 10−3). The observed mutational features for STRs have important consequences for forensic applications, including the definition of criteria for exclusion in paternity testing and the interpretation of DNA profiles in identification analysis.

Population genetic analyses of the AmpFlSTR ® NGM™ in Brazil

Population data of 15 short tandem repeat loci of the AmpFlSTR® next generation multiplex (NGM)™ were obtained from a sample of 835 individuals. The loci are the ten short tandem repeats (STRs) in the SGM Plus® Kit plus the EDNAP- and ENSFI-recommended STRs D10S1248, D22S1045, D2S441, D1S1656, and D12S391. Allele frequency and other forensically relevant statistics data were generated for the NGM loci into five current country macroregions of Brazil (North, Northeast, Central West, Southeast, and South). All the analyzed loci meet Hardy–Weinberg equilibrium expectations and no linkage disequilibrium in all pairs of loci. The observed and expected heterozygosity, power of discrimination, polymorphic information content, and the other population–genetic indices were calculated. The overall power of discrimination was greater than 0.99999999999999999996 and the combined power of exclusion was greater than 0.9999998 in all Brazilian populations. Comparative analysis between populations from different Brazilian macroregions as well as between Brazil and Caucasian, African Americans, and Hispanic US populations are presented.

15 STR loci frequencies with mutation rates in the population from Rio Grande do Sul, Southern Brazil

Allele frequencies for 15 short tandem repeats (STR) loci were obtained from a sample of 2038 individuals undergoing paternity testing. The population is from Rio Grande do Sul, Brazil. The loci are the most commonly used in forensic and paternity testing, being analysed by the AmpFlSTR Identifiler (Applied Biosystems) commercial kit. The most polymorphic locus was D2S1338. Mutation rates were ascertained from this population sample. All the loci analysed reached the Hardy-Weinberg equilibrium. The results of genetic distance are consistent with the European-derived origins of Rio Grande do Sul population.

Population genetic data for 11 STR loci, including SE33, in Southern Brazil

Allele frequencies for 11 STR autosomal loci (D3S1358, vWA, D16S539, D2S1338, D8S1179, SE33, D19S433, TH01, FGA, D21S11, D18S51) were analysed in unrelated individuals undergoing paternity testing in Rio Grande do Sul State, Southern Brazil. The most polimorphic locus was SE33. The distributions of the genotypes in the evaluated loci are in Hardy-Weinberg equilibrium after Bonferronis correction.

Association between cytokine gene polymorphisms and outcome of hepatitis B virus infection in southern Brazil

A number of studies have demonstrated associations between cytokine gene polymorphisms and outcome of hepatitis B virus (HBV) infection. However, no general consensus has been reached, possibly due to differences between ethnic groups. In this study, 345 individuals living in southern Brazil, including 196 chronic HBV carriers and 149 subjects who had spontaneously recovered from acute infection, were enrolled to evaluate the influence of cytokine gene polymorphisms on the outcome of HBV infection. Most participants were of European descent. Genotyping of IL2‐330 G/T, IL4‐589C/T, IL6‐174 G/C, IL10‐592C/A, IL10‐1082 A/G, IL17A‐197 G/A, IL17A‐692 T/C, TNF‐α‐238 G/A, and TNF‐α‐308 G/A single nucleotide polymorphisms was performed by using the minisequencing (single base extension) method. By multivariable analysis, a statistically significant association was found between genotypic profile AA + GA in TNF‐α‐308 and chronic HBV infection (OR, 1.82; 95%CI, 1.01–3.27; P = 0.046). In southern Brazil, the carriers of the −308A allele in the TNF‐α gene promoter have a moderately higher risk of becoming chronic carriers in case of HBV infection. In addition, patients with chronic active hepatitis B (n = 60) exhibited a decreased frequency (3.3%) of the TNF‐238A allele when compared to that (14.8%) found among asymptomatic HBV carriers (n = 136), suggesting that this could be a protective factor against liver injury (OR, 0.17; 95%CI, 0.04–0.076; P = 0.023). J. Med. Virol. 88:1759–1766, 2016.

15 STR loci frequencies in the population from Paraná, Southern Brazil

Allele frequencies for 15 short tandem repeats (STR) loci were obtained from a sample of 4076 unrelated individuals undergoing paternity testing. The population is from Paraná, Southern Brazil. The loci are the most commonly used in forensic and paternity testing, being analyzed by the AmpFlSTR((R)) Identifiler (Applied Biosystems) commercial kit. The most polymorphic loci were D2S1338 and D18S51. Excepting the D13S317, all loci were in Hardy-Weinberg equilibrium. Comparative analyses between our population data and other populations are presented.

Investigation of paternity with alleged father deceased or missing: Analysis of success at the end of the report

In this work we present a retrospective study of 858 cases of paternity investigation performed in Rio Grande do Sul, Southern Brazil, from 2007 to 2012, where the alleged father was deceased or missing. These cases represent 3.3% (858/26187) of paternity tests performed in that period. Considering the analysis of 17 DNA short tandem repeat loci, we present here the proportion of cases with conclusive results according to the number of relatives of the unavailable alleged father investigated and their kinship. The results show 81.0% (695/858) of cases with conclusive results and their characteristics.

Genetic diversity of Mycobacterium tuberculosis isoniazid monoresistant and multidrug-resistant in Rio Grande do Sul, a tuberculosis high-burden state in Brazil

Tuberculosis (TB) remains a major public health problem in the world and Brazil is among the countries with the highest incidence and prevalence rates, and Rio Grande do Sul, a Brazilian state, occupy a prominent position. Multidrug-resistant Mycobacterium tuberculosis (MDR-TB) further aggravates this scenario, making it more difficult to treat and control the disease. Isoniazid monoresistance (IMR) may increase the risk of progression to MDR-TB and treatment failure. However, most drug resistance molecular tests only focus on detecting rifampicin (RIF) resistance.In the present study, we characterized a total of 63 drug resistant isolates of M. tuberculosis (35 MDR, 26 IMR and two isolates monoresistant to rifampicin [RMR]) of the Rio Grande do Sul state by MIRU-VNTR (24 loci), spoligotyping, presence of RDRio, fbpC103, pks15/1 and sequencing of the katG, rpoB and inhA genes. We observed a higher proportion of the LAM family 30/63 (47.61%). In IMR, mutations were found in the katG gene (98% at codon 315) in 72.5%, and mutations in the promoter region of the inhA gene in 6.25% of the isolates. In MDR-TB and RMR-TB isolates, 92.1% had mutations in the rpoB gene (57% at codon 531). The presence of a 12 bp insertion between codons 516 and 517 of the rpoB gene in MDR-TB isolates was found in five isolates. In conclusion, we observed that the highest frequency of IMR-TB and MDR-TB strains belong to the LAM and Haarlem genotypes in Rio Grande do Sul state. A significant number of isolates previously characterized as Mycobacterium pinnipedi2 through spoligotyping were found to belong to the M. tuberculosis LAM family. This was responsible for a number of significant cases and the molecular profile of this strain and the pattern of mutations related to drug resistance were analyzed. These findings may contribute to a better understanding about the spread of M. tuberculosis resistant in southern of Brazil.

Genetic diagnosis in recently transfused patients.

Analysis of recently transfused patients is usually postponed to avoid spurious results because of contamination with donor’s cells. However, little is known about the extent of this influence in routine molecular diagnostic tests. To elucidate this question, we tested a mix of blood samples from 2 &agr;-1-antitrypsin-deficient patients diagnosed as Pi*Z homozygous with 1 normal donor at 1:1, 1:10, 1:20, and 1:30 proportions. Human identification panel and Pi*Z allele detection were used to establish the detection limit of a blood mixture. Mixtures of 1:1 and 1:10 were easily detected with both techniques, whereas for 1:30, it was necessary to change the equipment settings to identify the mixture. Moreover, the heterozygous pattern observed for the mixtures on Pi*Z genotyping was weaker at this level of mixture. We further evaluated the degree of mixture detectable in 20 transfused patients who received 1 blood unit (concentrate of irradiated or nonirradiated red blood cells) using the human identification panel. Two days after the transfusion, the presence of the donor’s markers was not detected, suggesting that after this time point the levels of admixture are below 1:30. The methods applied in the present study showed adequate sensitivity to identify alleles of the so-called “smaller population” of cells up to 3%, approximately. The same result was obtained in a “diagnostic situation,” in which the blood mixture was submitted to a PCR-RFLP protocol to detect a mutation.