RESUMO
Cyanobacterial blooms affect biotic interactions in aquatic ecosystems, including those involving heterotrophic bacteria. Ultra-small microbial communities are found in both surface water and groundwater and include diverse heterotrophic bacteria. Although the taxonomic composition of these communities has been described in some environments, the involvement of these small cells in the fate of environmentally relevant molecules has not been investigated. Here, we aimed to test if small-sized microbial fractions from a polluted urban lagoon were able to degrade the cyanotoxin microcystin (MC). We obtained cells after filtration through 0.45 as well as 0.22 µm membranes and characterized the morphology and taxonomic composition of bacteria before and after incubation with and without microcystin-LR (MC-LR). Communities from different size fractions (< 0.22 and < 0.45 µm) were able to remove the dissolved MC-LR. The originally small-sized cells grew during incubation, as shown by transmission electron microscopy, and changed in both cell size and morphology. The analysis of 16S rDNA sequences revealed that communities originated from < 0.22 and < 0.45 µm fractions diverged in taxonomic composition although they shared certain bacterial taxa. The presence of MC-LR shifted the structure of < 0.45 µm communities in comparison to those maintained without toxin. Actinobacteria was initially dominant and after incubation with MC-LR Proteobacteria predominated. There was a clear enhancement of taxa already known to degrade MC-LR such as Methylophilaceae. Small-sized bacteria constitute a diverse and underestimated fraction of microbial communities, which participate in the dynamics of MC-LR in natural environments.
Assuntos
Cianobactérias , Microbiota , Organismos Aquáticos , Cianobactérias/metabolismo , Filtração , Microcistinas/metabolismoRESUMO
Cyanobacteria stand out among phytoplankton when they form massive blooms and produce toxins. Because cyanotoxin genes date to the origin of metazoans, the hypothesis that cyanotoxins function as a defense against herbivory is still debated. Although their primary cellular function might vary, these metabolites could have evolved as an anti-predator response. Here we evaluated the physiological and molecular responses of a saxitoxin-producing Raphidiopsis raciborskii to infochemicals released by the grazer Daphnia gessneri. Induced chemical defenses were evidenced in R. raciborskii as a significant increase in the transcription level of sxt genes, followed by an increase in saxitoxin content when exposed to predator cues. Moreover, cyanobacterial growth decreased, and no significant effects on photosynthesis or morphology were observed. Overall, the induced defense response was accompanied by a trade-off between toxin production and growth. These results shed light on the mechanisms underlying zooplankton-cyanobacteria interactions in aquatic food webs. The widespread occurrence of the cyanobacterium R. raciborskii in freshwater bodies has been attributed to its phenotypic plasticity. Assessing the potential of this species to thrive over interaction filters such as zooplankton grazing pressure can enhance our understanding of its adaptive success.
Assuntos
Cylindrospermopsis , Daphnia/metabolismo , Feromônios/metabolismo , Saxitoxina , Zooplâncton/metabolismo , Animais , Cylindrospermopsis/genética , Cylindrospermopsis/crescimento & desenvolvimento , Cylindrospermopsis/metabolismo , Cadeia Alimentar , Regulação Bacteriana da Expressão Gênica , Genes Bacterianos , Saxitoxina/biossíntese , Saxitoxina/genéticaRESUMO
Phosphorus loading plays an important role in the occurrence of cyanobacterial blooms and understanding how this nutrient affects the physiology of cyanobacteria is imperative to manage these phenomena. Microcystis aeruginosa and Raphidiopsis raciborskii are cyanobacterial species that form potentially toxic blooms in freshwater ecosystems worldwide. Blooms comprise numerous strains with high trait variability, which can contribute to the widespread distribution of these species. Here, we explored the intraspecific variability in response to phosphorus depleted conditions (P-) testing five strains of each species. Strains could be differentiated by cell volume or genetic profiles except for those of the same species, sampling location and date, though these presented differences in their response to (P-). Although differently affected by (P-) over 10 days, all strains were able to grow and maintain photosynthetic activity. For most M. aeruginosa and R. raciborskii strains growth rates were not significantly different comparing (P+) and (P-) conditions. After ten days in (P-), only one M. aeruginosa strain and two R. raciborskii strains showed reduction in biovolume yield as compared to (P+) but in most strains chlorophyll-a concentrations were lower in (P-) than in (P+). Reduced photosystem II efficiency was found for only one R. raciborskii strain while all M. aeruginosa strains were affected. Only two M. aeruginosa and one R. raciborskii strain increased alkaline phosphatase activity under (P-) as compared to (P+). Variation in P-uptake was also observed but comparison among strains yielded homogeneous groups comprised of representatives of both species. Comparing the response of each species as a whole, the (P-) condition affected growth rate, biovolume yield and chlorophyll yield. However, these parameters revealed variation among strains of the same species to the extent that differences between M. aeruginosa and R. raciborskii were not significant. Taken together, these results do not support the idea that R. raciborskii, as a species, can withstand phosphorus limitation better than M. aeruginosa and also point that the level of intraspecific variation may preclude generalizations based on studies that use only one or few strains.
Assuntos
Cianobactérias , Cylindrospermopsis , Microcystis , Ecossistema , FósforoRESUMO
OBJECTIVE: The aim of this study was to investigate the proteome of the gingival crevicular fluid comparing the relative abundance of proteins from type 2 diabetes mellitus (2DM) individuals and chronic periodontitis (CP) affected sites, subjects affected by both conditions and healthy individuals. MATERIAL AND METHODS: Twenty individuals were equally allocated in four groups, 2DM with CP, 2DM periodontally healthy, CP without 2DM, and periodontally healthy without 2DM. The relative quantification of proteins was accessed with iTRAQ labeling and mass spectrometry. RESULTS AND CONCLUSION: A total of 104 proteins showed significant differences in abundance in pairwise comparisons. Some presented different levels in all diseased groups as compared to control, either increasing (rap guanine nucleotide exchange factor, S100A8, S100A9, and immunoglobulins) or decreasing (actins, myristoylated alanine-rich C-kinase substrate, and glutathione S-transferase). Other differences were specific for a given condition: Titin, neutrophil elastase, and myeloperoxidase levels were higher in the DP group, cathelicidin antimicrobial peptide decreased in CP, and annexin decreased in DH. These differences in the proteome can provide clues for further studies that will validate the variation in their levels and their role in both diseases.
Assuntos
Periodontite Crônica/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Líquido do Sulco Gengival/química , Proteoma/análise , Idoso , Estudos de Casos e Controles , Cromatografia Líquida , Periodontite Crônica/complicações , Diabetes Mellitus Tipo 2/complicações , Feminino , Humanos , Masculino , Espectrometria de Massas , Pessoa de Meia-IdadeRESUMO
The wide distribution of cyanobacteria in aquatic environments leads to the risk of water contamination by cyanotoxins, which generate environmental and public health issues. Measurements of cell densities or pigment contents allow both the early detection of cellular growth and bloom monitoring, but these methods are not sufficiently accurate to predict actual cyanobacterial risk. To quantify cyanotoxins, analytical methods are considered the gold standards, but they are laborious, expensive, time-consuming and available in a limited number of laboratories. In cyanobacterial species with toxic potential, cyanotoxin production is restricted to some strains, and blooms can contain varying proportions of both toxic and non-toxic cells, which are morphologically indistinguishable. The sequencing of cyanobacterial genomes led to the description of gene clusters responsible for cyanotoxin production, which paved the way for the use of these genes as targets for PCR and then quantitative PCR (qPCR). Thus, the quantification of cyanotoxin genes appeared as a new method for estimating the potential toxicity of blooms. This raises a question concerning whether qPCR-based methods would be a reliable indicator of toxin concentration in the environment. Here, we review studies that report the parallel detection of microcystin genes and microcystin concentrations in natural populations and also a smaller number of studies dedicated to cylindrospermopsin and saxitoxin. We discuss the possible issues associated with the contradictory findings reported to date, present methodological limitations and consider the use of qPCR as an indicator of cyanotoxin risk.
Assuntos
Toxinas Bacterianas/isolamento & purificação , Cianobactérias/crescimento & desenvolvimento , Monitoramento Ambiental/métodos , Água Doce/microbiologia , Reação em Cadeia da Polimerase em Tempo Real , Alcaloides , Toxinas Bacterianas/genética , Cianobactérias/genética , Toxinas de Cianobactérias , Proliferação Nociva de Algas , Microcistinas/genética , Microcistinas/isolamento & purificação , RNA Ribossômico 16S/genética , Saxitoxina/genética , Saxitoxina/isolamento & purificação , Uracila/análogos & derivados , Uracila/isolamento & purificaçãoRESUMO
Enterotoxigenic Escherichia coli (ETEC) strains have been implicated as important etiological agents of diarrheal disease, especially in developing countries. This group of microorganisms has been associated with a diverse range of genotypic and phenotypic markers. In the present study, 21 ETEC isolates previously defined according to the toxigenic genotypes, were characterized on the basis of O:H typing, cell adherence patterns, and colonization factors (CFs) antigens. Genetic diversity was investigated by random amplification polymorphic DNA (RAPD-PCR), pulsed-field gel electrophoresis (PFGE) and multilocus enzyme electrophoresis (MLEE). LT-I probe-positive isolates belonged to serotypes ONT:HNT, O7:H24, O48:H21, O88:H25, O148:H28, O159:H17 and O159:H21. ST-h probe-positive isolates belonged to serotypes O159:H17, O148:H28 and O6:H-. Serotypes O148:H28, O159:H17 and O6:H- were associated with the CS6, CFA/I and CS1 CS3 antigens, respectively. Most ETEC strains exhibited a diffuse pattern of adherence to cultured epithelial cells. In general, phenotypic and genotypic characteristics correlated well. RAPD-PCR, PFGE and MLEE showed reproducibility and good discriminatory potential. The application of molecular typing systems allowed the detection of significant diversity among the isolates, indicating a non-clonal origin and revealing intra-serotype variation overlooked by classical epidemiological approaches. The phenotypic and genotypic diversity observed lead us to recommend the use of different typing systems in order to elucidate the epidemiology of ETEC infection.
Assuntos
Infecções por Escherichia coli/microbiologia , Escherichia coli/classificação , Genótipo , Fenótipo , Brasil , Enterotoxinas/metabolismo , Escherichia coli/genética , Escherichia coli/isolamento & purificação , Proteínas de Fímbrias , Variação Genética , Filogenia , Técnica de Amplificação ao Acaso de DNA Polimórfico , SorotipagemRESUMO
O gene da RNA-polimerase do vírus da febre aftosa foi mutagenizado dentro do seu sítio de atividade. Utilizando o cDNA da cepa viral CS8 C1 - Santa Pau, o gene foi digerido com Pst I gerando um fragmento contendo a seqüência crítica de mutagênese. Este fragmento foi subclonado em M13mp8 e quatro mutaçöes foram geradas in vitro através do método de mutagênese sítio-dirigida por oligonucleotídeo. O gene da polimerase foi entäo reconstruído e subclonado em pUC19. Estes mutantes seräo usados em estudos de estrutura e atividade da enzima e no desenvolvimento da técnica de "imunizaçäo intracelular" em células eurcariontes