New insights on tuberculosis transmission dynamics and drug susceptibility profiles among the prison population in Southern Brazil based on whole-genome sequencing.

BACKGROUND: The rate of tuberculosis (TB) infection among the prison population (PP) in Brazil is 28 times higher than that in the general population, and prison environment favors the spread of TB. OBJECTIVE: To describe TB transmission dynamics and drug resistance profiles in PP using whole-genome sequencing (WGS). METHODS: This was a retrospective study of Mycobacterium tuberculosis cultivated from people incarcerated in 55 prisons between 2016 and 2019; only one isolate per prisoner was included. Information about movement from one prison to another was tracked. Clinical information was collected, and WGS was performed on isolates obtained at the time of TB diagnosis. RESULTS: Among 134 prisoners included in the study, we detected 16 clusters with a total of 58 (43%) cases of M. tuberculosis. Clusters ranged from two to seven isolates with five or fewer single nucleotide polymorphism (SNP) differences, suggesting a recent transmission. Six (4.4%) isolates were resistant to at least one anti-TB drug. Two of these clustered together and showed resistance to rifampicin, isoniazid, and fluoroquinolones, with 100% concordance between WGS and phenotypic drug-susceptibility testing. Prisoners with clustered isolates had a high amount of movement between prisons (two to eight moves) during the study period. CONCLUSIONS: WGS demonstrated the recent transmission of TB within prisons in Brazil. The high movement among prisoners seems to be related to the transmission of the same M. tuberculosis strain within the prison system. Screening for TB before and after the movement of prisoners using rapid molecular tests could play a role in reducing transmission.

New insights on tuberculosis transmission dynamics and drug susceptibility profiles among the prison population in Southern Brazil based on whole-genome sequencing

ABSTRACT Background: The rate of tuberculosis (TB) infection among the prison population (PP) in Brazil is 28 times higher than that in the general population, and prison environment favors the spread of TB. Objective: To describe TB transmission dynamics and drug resistance profiles in PP using whole-genome sequencing (WGS). Methods: This was a retrospective study of Mycobacterium tuberculosis cultivated from people incarcerated in 55 prisons between 2016 and 2019; only one isolate per prisoner was included. Information about movement from one prison to another was tracked. Clinical information was collected, and WGS was performed on isolates obtained at the time of TB diagnosis. Results: Among 134 prisoners included in the study, we detected 16 clusters with a total of 58 (43%) cases of M. tuberculosis. Clusters ranged from two to seven isolates with five or fewer single nucleotide polymorphism (SNP) differences, suggesting a recent transmission. Six (4.4%) isolates were resistant to at least one anti-TB drug. Two of these clustered together and showed resistance to rifampicin, isoniazid, and fluoroquinolones, with 100% concordance between WGS and phenotypic drug-susceptibility testing. Prisoners with clustered isolates had a high amount of movement between prisons (two to eight moves) during the study period. Conclusions: WGS demonstrated the recent transmission of TB within prisons in Brazil. The high movement among prisoners seems to be related to the transmission of the same M. tuberculosis strain within the prison system. Screening for TB before and after the movement of prisoners using rapid molecular tests could play a role in reducing transmission.

Germline variants in DNA repair genes associated with hereditary breast and ovarian cancer syndrome: analysis of a 21 gene panel in the Brazilian population.

BACKGROUND: The Hereditary Breast and Ovarian Cancer Syndrome (HBOC) occurs in families with a history of breast/ovarian cancer, presenting an autosomal dominant inheritance pattern. BRCA1 and BRCA2 are high penetrance genes associated with an increased risk of up to 20-fold for breast and ovarian cancer. However, only 20-30% of HBOC cases present pathogenic variants in those genes, and other DNA repair genes have emerged as increasing the risk for HBOC. In Brazil, variants in ATM, ATR, CHEK2, MLH1, MSH2, MSH6, POLQ, PTEN, and TP53 genes have been reported in up to 7.35% of the studied cases. Here we screened and characterized variants in 21 DNA repair genes in HBOC patients. METHODS: We systematically analyzed 708 amplicons encompassing the coding and flanking regions of 21 genes related to DNA repair pathways (ABRAXAS1, ATM, ATR, BARD1, BRCA1, BRCA2, BRIP1, CDH1, CHEK2, MLH1, MRE11, MSH2, MSH6, NBN, PALB2, PMS2, PTEN, RAD50, RAD51, TP53 and UIMC1). A total of 95 individuals with HBOC syndrome clinical suspicion in Southeast Brazil were sequenced, and 25 samples were evaluated for insertions/deletions in BRCA1/BRCA2 genes. Identified variants were assessed in terms of population allele frequency and their functional effects were predicted through in silico algorithms. RESULTS: We identified 80 variants in 19 genes. About 23.4% of the patients presented pathogenic variants in BRCA1, BRCA2 and TP53, a frequency higher than that identified among previous studies in Brazil. We identified a novel variant in ATR, which was predicted as pathogenic by in silico tools. The association analysis revealed 13 missense variants in ABRAXAS1, BARD1, BRCA2, CHEK2, CDH1, MLH1, PALB2, and PMS2 genes, as significantly associated with increased risk to HBOC, and the patients carrying those variants did not present large insertions or deletions in BRCA1/BRCA2 genes. CONCLUSIONS: This study embodies the third report of a multi-gene analysis in the Brazilian population, and addresses the first report of many germline variants associated with HBOC in Brazil. Although further functional analyses are necessary to better characterize the contribution of those variants to the phenotype, these findings would improve the risk estimation and clinical follow-up of patients with HBOC clinical suspicion.

Comparative transcriptomics in Leishmania braziliensis: disclosing differential gene expression of coding and putative noncoding RNAs across developmental stages.

Leishmaniasis is a worldwide public health problem caused by protozoan parasites of the genus Leishmania. Leishmania braziliensis is the most important species responsible for tegumentary leishmaniases in Brazil. An understanding of the molecular mechanisms underlying the success of this parasite is urgently needed. An in-depth study on the modulation of gene expression across the life cycle stages of L. braziliensis covering coding and noncoding RNAs (ncRNAs) was missing and is presented herein. Analyses of differentially expressed (DE) genes revealed that most prominent differences were observed between the transcriptomes of insect and mammalian proliferative forms (6,576 genes). Gene ontology (GO) analysis indicated stage-specific enriched biological processes. A computational pipeline and 5 ncRNA predictors allowed the identification of 11,372 putative ncRNAs. Most of the DE ncRNAs were found between the transcriptomes of insect and mammalian proliferative stages (38%). Of the DE ncRNAs, 295 were DE in all three stages and displayed a wide range of lengths, chromosomal distributions and locations; many of them had a distinct expression profile compared to that of their protein-coding neighbors. Thirty-five putative ncRNAs were submitted to northern blotting analysis, and one or more hybridization-positive signals were observed in 22 of these ncRNAs. This work presents an overview of the L. braziliensis transcriptome and its adjustments throughout development. In addition to determining the general features of the transcriptome at each life stage and the profile of protein-coding transcripts, we identified and characterized a variety of noncoding transcripts. The novel putative ncRNAs uncovered in L. braziliensis might be regulatory elements to be further investigated.

Assessing the efficiency of multiple sequence alignment programs.

BACKGROUND: Multiple sequence alignment (MSA) is an extremely useful tool for molecular and evolutionary biology and there are several programs and algorithms available for this purpose. Although previous studies have compared the alignment accuracy of different MSA programs, their computational time and memory usage have not been systematically evaluated. Given the unprecedented amount of data produced by next generation deep sequencing platforms, and increasing demand for large-scale data analysis, it is imperative to optimize the application of software. Therefore, a balance between alignment accuracy and computational cost has become a critical indicator of the most suitable MSA program. We compared both accuracy and cost of nine popular MSA programs, namely CLUSTALW, CLUSTAL OMEGA, DIALIGN-TX, MAFFT, MUSCLE, POA, Probalign, Probcons and T-Coffee, against the benchmark alignment dataset BAliBASE and discuss the relevance of some implementations embedded in each program's algorithm. Accuracy of alignment was calculated with the two standard scoring functions provided by BAliBASE, the sum-of-pairs and total-column scores, and computational costs were determined by collecting peak memory usage and time of execution. RESULTS: Our results indicate that mostly the consistency-based programs Probcons, T-Coffee, Probalign and MAFFT outperformed the other programs in accuracy. Whenever sequences with large N/C terminal extensions were present in the BAliBASE suite, Probalign, MAFFT and also CLUSTAL OMEGA outperformed Probcons and T-Coffee. The drawback of these programs is that they are more memory-greedy and slower than POA, CLUSTALW, DIALIGN-TX, and MUSCLE. CLUSTALW and MUSCLE were the fastest programs, being CLUSTALW the least RAM memory demanding program. CONCLUSIONS: Based on the results presented herein, all four programs Probcons, T-Coffee, Probalign and MAFFT are well recommended for better accuracy of multiple sequence alignments. T-Coffee and recent versions of MAFFT can deliver faster and reliable alignments, which are specially suited for larger datasets than those encountered in the BAliBASE suite, if multi-core computers are available. In fact, parallelization of alignments for multi-core computers should probably be addressed by more programs in a near future, which will certainly improve performance significantly.

Complete genome sequence of type strain Campylobacter fetus subsp. venerealis NCTC 10354T.

Campylobacter fetus subsp. venerealis is the etiologic agent of bovine genital campylobacteriosis, a sexually transmitted disease of cattle that is of worldwide importance. The complete sequencing and annotation of the genome of the type strain C. fetus subsp. venerealis NCTC 10354(T) are reported.

Análise computacional baseada no desenvolvimento de um pipeline de técnicas ab initio para predição de desordem estrutural protéica em genomas de tripanosomatídeos / Computational analysis based on developing a pipeline of techniques for ab initio prediction of protein structural disorder in the genomes of trypanosomatids

Proteínas são compostas por uma ou mais cadeias de aminoácidos e exibem vários níveis de organização estrutural. Recentemente, uma classe de proteínas conhecidas como IUPs (Intrinsically Unstructured Proteins) foi descoberta e sua principal característica é a ausência de estrutura parcial ou total em seu estado nativo. Devido à sua adaptabilidade intrínseca, tais proteínas participam em muitos. processos biológicos regulatórios incluindo o escape do sistema imune. Utilizando a informação contida no proteoma predito de Leishmania braziliensis, Leishmania major, Leishmania infantum, Trypanosoma cruzi e Trypanosoma brucei, desenvolvemos um pipeline de análise computacional que tem como objetivo a identificação, caracterização e análise de IUPs. Nosso principal objetivo é investigar. as correlações biológicas entre desordem estrutural e as interações parasitohospedeiro.. O pipeline emprega 6 metodologias de predição de desordem, integra informações obtidas através da anotação estrutural e funcional, predição subcelular e o cálculo de propriedades físico-químicas. Como cerne do pipeline de IUPs existe um banco de dados relacional modelado de forma a viabilizar a extração das relações entre as inúmeras variáveis analisadas e gerar os relatórios.

Nossos resultados demonstram que as espécies de Leishmania e Trypanosoma possuem aproximadamente 70% e 55% de IUPs, respectivamente. Nossos resultados indicam que nos tripanosomatídeos os aminoácidos promotores de ordem são: W, Y, F, V, I, L e C e os promotores de desordem são P, Q, E, R e S.. A anotação funcional revelou o enriquecimento dos termos GO: transcription, ribonucleoproteins, RNA metabolic process, protein binding e ribonucleotide binding.. Outra característica é a associação entre o aumento da porcentagem de resíduos desordenados, a localização subcelular e o número de regiões transmembrana. Como resultado da validação experimental feita através de técnicas de eletroforese bidimensional (2D) específicas para identificação de IUPs, 100% dos spots identificados foram preditos in silico.. Até o presente momento este trabalho representa a primeira tentativa de estabelecimento das correlações entre desordem estrutural protéica e função nos tripanosomatídeos. A execução do pipeline pode ser solicitada por instituições acadêmicas através de solicitação no sitio http://iup.cpqrr.fiocruz.br/iup-pipeline.

Análise computacional baseada no desenvolvimento de um pipeline de técnicas ab initio para predição de desordem estrutural protéica em genomas de tripanosomatídeos

Proteínas são compostas por uma ou mais cadeias de aminoácidos e exibem vários níveis de organização estrutural. Recentemente, uma classe de proteínas conhecidas como IUPs (Intrinsically Unstructured Proteins) foi descoberta e sua principal característica é a ausência de estrutura parcial ou total em seu estado nativo. Devido à sua adaptabilidade intrínseca, tais proteínas participam em muitos. processos biológicos regulatórios incluindo o escape do sistema imune. Utilizando a informação contida no proteoma predito de Leishmania braziliensis, Leishmania major, Leishmania infantum, Trypanosoma cruzi e Trypanosoma brucei, desenvolvemos um pipeline de análise computacional que tem como objetivo a identificação, caracterização e análise de IUPs. Nosso principal objetivo é investigar. as correlações biológicas entre desordem estrutural e as interações parasitohospedeiro.. O pipeline emprega 6 metodologias de predição de desordem, integra informações obtidas através da anotação estrutural e funcional, predição subcelular e o cálculo de propriedades físico-químicas. Como cerne do pipeline de IUPs existe um banco de dados relacional modelado de forma a viabilizar a extração das relações entre as inúmeras variáveis analisadas e gerar os relatórios.

Nossos resultados demonstram que as espécies de Leishmania e Trypanosoma possuem aproximadamente 70% e 55% de IUPs, respectivamente. Nossos resultados indicam que nos tripanosomatídeos os aminoácidos promotores de ordem são: W, Y, F, V, I, L e C e os promotores de desordem são P, Q, E, R e S.. A anotação funcional revelou o enriquecimento dos termos GO: transcription, ribonucleoproteins, RNA metabolic process, protein binding e ribonucleotide binding.. Outra característica é a associação entre o aumento da porcentagem de resíduos desordenados, a localização subcelular e o número de regiões transmembrana. Como resultado da validação experimental feita através de técnicas de eletroforese bidimensional (2D) específicas para identificação de IUPs, 100% dos spots identificados foram preditos in silico.. Até o presente momento este trabalho representa a primeira tentativa de estabelecimento das correlações entre desordem estrutural protéica e função nos tripanosomatídeos. A execução do pipeline pode ser solicitada por instituições acadêmicas através de solicitação no sitio http://iup.cpqrr.fiocruz.br/iup-pipeline.