Genome analysis of DNA repair genes in the alpha proteobacterium Caulobacter crescentus

BACKGROUND: The integrity of DNA molecules is fundamental for maintaining life. The DNA repair proteins protect organisms against genetic damage, by removal of DNA lesions or helping to tolerate them. DNA repair genes are best known from the gamma-proteobacterium Escherichia coli, which is the most understood bacterial model. However, genome sequencing raises questions regarding uniformity and ubiquity of these DNA repair genes and pathways, reinforcing the need for identifying genes and proteins, which may respond to DNA damage in other bacteria. RESULTS: In this study, we employed a bioinformatic approach, to analyse and describe the open reading frames potentially related to DNA repair from the genome of the alpha-proteobacterium Caulobacter crescentus. This was performed by comparison with known DNA repair related genes found in public databases. As expected, although C. crescentus and E. coli bacteria belong to separate phylogenetic groups, many of their DNA repair genes are very similar. However, some important DNA repair genes are absent in the C. crescentus genome and other interesting functionally related gene duplications are present, which do not occur in E. coli. These include DNA ligases, exonuclease III (xthA), endonuclease III (nth), O(6)-methylguanine-DNA methyltransferase (ada gene), photolyase-like genes, and uracil-DNA-glycosylases. On the other hand, the genes imuA and imuB, which are involved in DNA damage induced mutagenesis, have recently been described in C. crescentus, but are absent in E. coli. Particularly interesting are the potential atypical phylogeny of one of the photolyase genes in alpha-proteobacteria, indicating an origin by horizontal transfer, and the duplication of the Ada orthologs, which have diverse structural configurations, including one that is still unique for C. crescentus. CONCLUSION: The absence and the presence of certain genes are discussed and predictions are made considering the particular aspects of the C. crescentus among other known DNA repair pathways. The observed differences enlarge what is known for DNA repair in the Bacterial world, and provide a useful framework for further experimental studies in this organism

Different patterns of evolution for duplicated DNA repair genes in bacteria of the Xanthomonadales group

BACKGROUND: DNA repair genes encode proteins that protect organisms against genetic damage generated by environmental agents and by-products of cell metabolism. The importance of these genes in life maintenance is supported by their high conservation, and the presence of duplications of such genes may be easily traced, especially in prokaryotic genomes. RESULTS: The genome sequences of two Xanthomonas species were used as the basis for phylogenetic analyses of genes related to DNA repair that were found duplicated. Although 16S rRNA phylogenetic analyses confirm their classification at the basis of the gamma proteobacteria subdivision, differences were found in the origin of the various genes investigated. Except for lexA, detected as a recent duplication, most of the genes in more than one copy are represented by two highly divergent orthologs. Basically, one of such duplications is frequently positioned close to other gamma proteobacteria, but the second is often positioned close to unrelated bacteria. These orthologs may have occurred from old duplication events, followed by extensive gene loss, or were originated from lateral gene transfer (LGT), as is the case of the uvrD homolog. CONCLUSIONS: Duplications of DNA repair related genes may result in redundancy and also improve the organisms' responses to environmental challenges. Most of such duplications, in Xanthomonas, seem to have arisen from old events and possibly enlarge both functional and evolutionary genome potentiality

Overexpression of KLC2 due to a homozygous deletion in the non-coding region causes SPOAN syndrome

SPOAN syndrome is a neurodegenerative disorder mainly characterized by spastic paraplegia, optic atrophy and neuropathy (SPOAN). Affected patients are wheelchair bound after 15 years old, with progressive joint contractures and spine deformities. SPOAN patients also have sub normal vision secondary to apparently non-progressive congenital optic atrophy. A potential causative gene was mapped at 11q13 ten years ago. Here we performed next-generation sequencing in SPOAN-derived samples. While whole-exome sequencing failed to identify the causative mutation, whole-genome sequencing allowed to detect a homozygous 216-bp deletion (chr11.hg19:g.66,024,557_66,024,773del) located at the non-coding upstream region of the KLC2 gene. Expression assays performed with patient’s fibroblasts and motor neurons derived from SPOAN patients showed KLC2 overexpression. Luciferase assay in constructs with 216-bp deletion confirmed the overexpression of gene reporter, varying from 48 to 74%, as compared with wild-type. Knockdown and overexpression of klc2 in Danio rerio revealed mild to severe curly-tail phenotype, which is suggestive of a neuromuscular disorder. Overexpression of a gene caused by a small deletion in the non-coding region is a novel mechanism, which to the best of our knowledge, was never reported before in a recessive condition. Although the molecular mechanism of KLC2 up-regulation still remains to be uncovered, such example adds to the importance of non-coding regions in human pathologyFil: Melo, Uira S.. Universidade de Sao Paulo; BrasilFil: Macedo Souza, Lucia I.. Universidade de Sao Paulo; BrasilFil: Figueiredo, Thalita. Federal University of Paraiba; Brasil. Paraiba State University; BrasilFil: Muotri, Alysson R. University of California at San Diego; Estados UnidosFil: Gleeson, Joseph G.. The Rockefeller University; Estados UnidosFil: Coux, Gabriela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: Armas, Pablo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: Calcaterra, Nora Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: Kitajima, João P.. Mendelics Genomic Analysis; BrasilFil: Amorim, Simone. Universidade de Sao Paulo; BrasilFil: Olávio, Thiago R.. Universidade de Sao Paulo; BrasilFil: Griesi Oliveira, Karina. Universidade de Sao Paulo; BrasilFil: Coatti, Giuliana C.. Universidade de Sao Paulo; BrasilFil: Rocha, Clarissa R.R. Universidade de Sao Paulo; BrasilFil: Martins Pinheiro, Marinalva. Universidade de Sao Paulo; BrasilFil: Menck, Carlos F.M.. Universidade de Sao Paulo; BrasilFil: Zaki, Maha S.. National Research Center. EL Cairo; EgiptoFil: Kok, Fernando. Universidade de Sao Paulo; BrasilFil: Zatz, Mayana. Universidade de Sao Paulo; BrasilFil: Santos, Silvana. Federal University of Paraiba; Brasil. Paraiba State University; Brasi

Search for genes related to the mutator phenotypes in Caulobacter crescentus.

A comparação \"in silico\" das vias de reparo de DNA nos genomas de Caulobacter crescentus e E. coli mostra diferenças significativas entre estas duas bactérias, sugerindo diversidade biológica das respostas a danos no DNA entre as bactérias. Em busca de genes que possam proteger o genoma de C. crescentus contra mutações, foi feita uma varredura em uma biblioteca de cerca de 5.000 clones construída a partir de inserção aleatória do transposon TN5 no genoma dessa bactéria. A maioria destes genes não tinha sido, ainda, relacionada a fenótipo mutador, mas, como esperado, também identificamos genes já conhecidos como relacionados a fenótipos mutadores. Alguns clones candidatos foram investigados quanto ao tipo de mutação induzida, baseando-se na sequência do gene rpoB, com o objetivo de indicar o processo mutacional sob condições genéticas específicas. Análises baseadas na medida da atividade promotora em fusão de transcrição com lacZ, mostra que o sistema SOS está alterado em alguns clones, e pode justificar parte do fenótipo mutador em alguns deles.The \"in silico\" comparison of the main DNA repair genes between C. crescentus and E. coli shows significant differences between these bacteria, suggesting biological diversity in bacterial responses to DNA damage. We further screened a C. crescentus library of 5,000 clones mutated by random insertion of the TN5 transposon in the genome, searching for clones with high levels of spontaneous rifampicin resistance mutations. Most of the genes identified have not been previously reported as related to mutator phenotype, but, as expected, we have also identified proteins already known as causing mutator phenotypes when mutated. As part of the functional characterization, some candidate clones were characterized based on rpoB gene sequences aiming at indicating the mutational process under specific genetic background conditions. Analysis of some candidate clones based on measurements of promoter activity with lacZ transcriptional fusions show that the system SOS is modified in some clones and this justify part of their mutator phenotype

Evolutionary and Functional Relationships of the dha Regulon by Genomic Context Analysis.

3-hydroxypropionaldehyde (3-HPA) and 1,3-propanediol (1,3-PD) are subproducts of glycerol degradation and of economical interest as they are used for polymers synthesis, such as polyesters and polyurethanes. Some few characterized bacterial species (mostly from Firmicutes and Gamma-proteobacteria groups) are able to catabolize these monomers from glycerol using the gene products from the dha regulon. To expand our knowledge and direct further experimental studies on the regulon and related genes for the anaerobic glycerol metabolism, an extensive genomic screening was performed to identify the presence of the dha genes in fully sequenced prokaryotic genomes. Interestingly, this work shows that although only few bacteria species are known to produce 3-HPA or 1,3-PD, the incomplete regulon is found in more than 100 prokaryotic genomes. However, the complete pathway is found only in a few dozen species belonging to five different taxonomic groups, including one Archaea species, Halalkalicoccus jeotgali. Phylogenetic analysis and conservation of both gene synteny and primary sequence similarity reinforce the idea that these genes have a common origin and were possibly acquired by lateral gene transfer (LGT). Besides the evolutionary aspect, the identification of homologs from several different organisms may predict potential alternative targets for faster or more efficient biological synthesis of 3-HPA or 1,3-PD

Qualidade Microbiológica do Leite in natura comercializado na cidade de Castro Alves-BA

Introdução: O leite é um alimento com excepcional valor nutritivo, amplamente consumido pela população mundial, entretanto, é também um excelente meio de cultura para muitos micro-organismos. A existência de problemas relacionados às condições higiênicas deficientes durante os processos de obtenção, manipulação e conservação são as principais razões para a perda de qualidade do leite. Objetivo: Considerando a importância que o leite assume na alimentação humana, realizou-se o presente trabalho com o objetivo de avaliar a qualidade microbiológica do leite cru, obtido por ordenha manual, comercializado na cidade de Castro Alves, Bahia, e comparar os resultados com os valores estabelecidos pela RDC Nº 12 que regulamenta os padrões microbiológicos para alimentos. Metodologia: Foram analisadas 20 amostras de leite cru, de janeiro a maio de 2011. Após a recepção das amostras junto aos comerciantes da região, estas foram acondicionadas, sob-refrigeração, e encaminhadas ao Laboratório de Microbiologia de Alimentos para determinação dos coliformes totais e fecais, isolamento de Escherichia coli, contagem de Staphylococcus aureus  e detecção direta de Salmonella. Resultados: Todas as amostras analisadas (100%) apresentaram valores elevados de coliformes totais, e coliformes fecais acima dos preconizados pela legislação. Seis amostras (30%) confirmaram a presença de E. coli, um importante enteropatógeno de origem fecal. Também foi evidenciado que 6 amostras estavam contaminadas com S.aureus. Salmonella spp. foi encontrada em 5 amostras (25%), micro-organismo esse que deve estar ausente em qualquer alimento destinado ao consumo humano. Conclusão: Os resultados encontrados nesta pesquisa demonstraram que a qualidade microbiológica do leite analisado esta insatisfatória, uma vez que nenhuma amostra atendeu aos requisitos estabelecidos pela legislação, principalmente devido a elevada contagem de coliformes fecais e presença de Salmonella spp. Tais achados constituem-se um motivo de preocupação em saúde pública, sobretudo, em função do risco representado pelo consumo desse produto pela população. Microbiological Quality of raw milk marketed in Castro Alves-BA Background: Milk is a food of exceptional nutritional value, widely consumed by the population; however, is also an excellent culture medium for many microorganisms. The existence of problems related to poor hygienic conditions during the process of obtaining, handling and storage are the main reasons for the loss of milk quality. Objective: Considering the importance that milk takes in food, took place this work in order to assess the microbiological quality of raw milk obtained by milking, marketed in the city of Castro Alves, Bahia, and compare the results with the values   established by RDC No. 12, which regulates the microbiological standards for foods. Methodology: We analyzed 20 samples of raw milk, from January to May 2011. After acquiring the samples along with local businessmen, they were placed under-cooling, and sent to the Food Microbiology Laboratory to determination of total and fecal coliforms, Escherichia coli isolation, Staphylococcus aureus on plates and direct detection of Salmonella. Results: All samples (100%) showed elevated levels of total coliforms and fecal coliforms above those recommended by the legislation. Six samples (30%) confirmed the presence of E. coli, an important enteropathogen of fecal origin. It was also shown that six samples were contaminated with S. aureus. Salmonella spp. was found in 5 samples (25%), the microorganism should be absent in other foods for human consumption. Conclusion: The findings of this study demonstrated that the microbiological quality of milk is considered unsatisfactory, since no sample meets the requirements established by legislation, mainly due to high fecal coliform counts and the presence of Salmonella spp.. These findings constitute a cause for concern in public health, mainly due to the risk posed by consumption of this product by the population. Keywords: Dairy Products. Quality Control.Coliforms. Pathogens

Genes identified for the reductive and oxidative pathway in the anaerobic metabolism of glycerol.

<p>Comparison of genes from <i>Klebsiella pneumoniae</i> to <i>B</i>.<i>intermedia</i>, <i>D</i>. <i>alkenivorans</i>, <i>H</i>. <i>jeotgali</i>, <i>Hyphomicrobium sp</i>., <i>I</i>. <i>polytropus</i>, <i>M</i>. <i>loti</i> and <i>M</i>. <i>opportunistum</i>.</p