RESUMO
In this work, the effect of different immobilization procedures on the properties of a lipase obtained from the extremophilic microorganism Serratia sp. USBA-GBX-513, which was isolated from Paramo soils of Los Nevados National Natural Park (Colombia), is reported. Different Shepharose beads were used: octyl-(OC), octyl-glyoxyl-(OC-GLX), cyanogen bromide (BrCN)-, and Q-Sepharose. The performance of the different immobilized extremophile lipase from Serratia (ESL) was compared with that of the lipase B from Candida antarctica (CALB). In all immobilization tests, hyperactivation of ESL was observed. The highest hyperactivation (10.3) was obtained by immobilization on the OC support. Subsequently, the thermal stability at pH 5, 7, and 9 and the stability in the presence of 50% (v/v) acetonitrile, 50% dioxane, and 50% tetrahydrofuran solvents at pH 7 and 40 °C were evaluated. ESL immobilized on octyl-Sepharose was the most stable biocatalyst at 90 °C and pH 9, while the most stable preparation at pH 5 was ESL immobilized on OC-GLX-Sepharose supports. Finally, in the presence of 50% (v/v) tetrahydrofuran (THF) or dioxane at 40 °C, ESL immobilized on OC-Sepharose was the most stable biocatalyst, while the immobilized preparation of ESL on Q-Sepharose was the most stable one in 40% (v/v) acetonitrile.
Assuntos
Proteínas de Bactérias/metabolismo , Enzimas Imobilizadas/metabolismo , Extremófilos/enzimologia , Lipase/metabolismo , Serratia/enzimologia , Basidiomycota/enzimologia , Biocatálise , Estabilidade Enzimática , Proteínas Fúngicas/metabolismo , Temperatura Alta , Concentração de Íons de Hidrogênio , Sefarose/análogos & derivados , Sefarose/químicaRESUMO
Serratia fonticola is a human pathogen widely found in the environment, with birds being reported as possible natural hosts. During an epidemiological and genomic surveillance study conducted to monitor the occurrence of extended-spectrum ß-lactamase (ESBL)-producing Enterobacterales in South American wild birds, we identified an ESBL-positive S. fonticola in a fecal sample collected from a Hudsonian Whimbrel, during its non-breeding range on the Pacific Coast of Chile. Whole genome sequencing analysis and "in silico" modeling revealed a novel variant of the class A ESBLs FONA family, designated FONA-7, which shows 96.28% amino acid identity with FONA-6; with amino acid substitutions occurring in the signal peptide sequence (Thr22âSer), and in the mature protein (Ser39âAsn and Thr227âIle). This finding denotes that migratory birds can be potential vectors for the transboundary spread of ESBL-producing bacteria, creating a further theoretical risk for the origin of novel plasmid-encoded ß-lactamases.
Assuntos
Antibacterianos/farmacologia , Proteínas de Bactérias/genética , Doenças das Aves/microbiologia , Serratia/efeitos dos fármacos , beta-Lactamases/genética , Animais , Aves/microbiologia , Chile/epidemiologia , Vetores de Doenças , Fezes , Polimorfismo Genético , Serratia/enzimologia , Serratia/genética , Sequenciamento Completo do GenomaRESUMO
Nine bacterial strains were previously isolated in association with pinewood nematode (PWN) from wilted pine trees. They proved to be nematicidal in vitro, and one of the highest activities, with potential to control PWN, was showed by Serratia sp. M24T3. Its ecology in association with plants remains unclear. This study aimed to evaluate the ability of strain M24T3 to colonize the internal tissues of the model plant Arabidopsis thaliana using confocal microscopy. Plant growth-promoting bacteria (PGPB) functional traits were tested and retrieved in the genome of strain M24T3. In greenhouse conditions, the bacterial effects of all nematicidal strains were also evaluated, co-inoculated or not with Bradyrhizobium sp. 3267, on Vigna unguiculata fitness. Inoculation of strain M24T3 increased the number of A. thaliana lateral roots and the confocal analysis confirmed effective bacterial colonization in the plant. Strain M24T3 showed cellulolytic activity, siderophores production, phosphate and zinc solubilization ability, and indole acetic acid production independent of supplementation with L-tryptophan. In the genome of strain M24T3, genes involved in the interaction with the plants such as 1-aminocyclopropane-1-carboxylate (ACC) deaminase, chitinolytic activity, and quorum sensing were also detected. The genomic organization showed ACC deaminase and its leucine-responsive transcriptional regulator, and the activity of ACC deaminase was 594.6 nmol α-ketobutyrate µg protein-1 µl-1. Strain M24T3 in co-inoculation with Bradyrhizobium sp. 3267 promoted the growth of V. unguiculata. In conclusion, this study demonstrated the ability of strain M24T3 to colonize other plants besides pine trees as an endophyte and displays PGPB traits that probably increased plant tolerance to stresses.
Assuntos
Arabidopsis/microbiologia , Nematoides/microbiologia , Serratia/fisiologia , Animais , Antibiose , Arabidopsis/crescimento & desenvolvimento , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Carbono-Carbono Liases/genética , Carbono-Carbono Liases/metabolismo , Pinus/parasitologia , Doenças das Plantas/parasitologia , Raízes de Plantas/crescimento & desenvolvimento , Raízes de Plantas/microbiologia , Percepção de Quorum , Serratia/enzimologia , Serratia/genética , Serratia/isolamento & purificação , Vigna/crescimento & desenvolvimento , Vigna/microbiologiaRESUMO
Ethylene acts as an inhibitor of the nodulation process of leguminous plants. However, some bacteria can decrease deleterious ethylene levels by the action of the enzyme 1-aminocyclopropane-1-carboxylate (ACC) deaminase which degrades ACC, the ethylene precursor in all higher plants. Co-inoculation of rhizobia with endophytes enhances the rhizobial symbiotic efficiency with legumes, improving both nodulation and nitrogen fixation. However, not much is understood about the mechanisms employed by these endophytic bacteria. In this regard, the role of ACC deaminase from endophytic strains in assisting rhizobia in this process has yet to be confirmed. In this study, the role of ACC deaminase in an endophyte's ability to increase Rhizobium tropici nodulation of common bean was evaluated. To assess the effect of ACC deaminase in an endophyte's ability to promote rhizobial nodulation, the endophyte Serratia grimesii BXF1, which does not encode ACC deaminase, was transformed with an exogenous acdS gene. The results obtained indicate that the ACC deaminase-overexpressing transformant strain increased common bean growth, and enhanced the nodulation abilities of R. tropici CIAT899, in both cases compared to the wild-type non-transformed strain. Furthermore, plant inoculation with the ACC deaminase-overproducing strain led to an increased level of plant protection against a seed-borne pathogen. SIGNIFICANCE AND IMPACT OF THE STUDY: In this work, we studied the effect of ACC deaminase production by the bacterial endophyte Serratia grimesi BXF1, and its impact on the nodulation process of common bean. The results obtained indicate that ACC deaminase is an asset to the synergetic interaction between rhizobia and the endophyte, positively contributing to the overall legume-rhizobia symbiosis by regulating inhibitory ethylene levels that might otherwise inhibit nodulation and overall plant growth. The use of rhizobia together with an ACC deaminase-producing endophyte is, therefore, an important strategy for the development of new bacterial inoculants with increased performance.
Assuntos
Proteínas de Bactérias/metabolismo , Carbono-Carbono Liases/metabolismo , Phaseolus/crescimento & desenvolvimento , Nodulação/fisiologia , Rhizobium tropici/metabolismo , Nódulos Radiculares de Plantas/microbiologia , Serratia/enzimologia , Inoculantes Agrícolas , Endófitos/metabolismo , Etilenos/metabolismo , Phaseolus/microbiologia , Serratia/genética , Serratia/metabolismo , SimbioseRESUMO
Carbapenemase-producing bacteria cause difficult-to-treat infections related to increased mortality in health care settings. Their occurrence has been reported in raw sewage, sewage-impacted rivers, and polluted coastal waters, which may indicate their spread to the community. We assessed the variety and concentration of carbapenemase producers in coastal waters with distinct pollution levels for 1 year. We describe various bacterial species producing distinct carbapenemases not only in unsuitable waters but also in waters considered suitable for primary contact.
Assuntos
Proteínas de Bactérias/genética , Klebsiella pneumoniae/genética , Água do Mar/microbiologia , Microbiologia da Água , beta-Lactamases/genética , Acinetobacter/enzimologia , Acinetobacter/genética , Acinetobacter/isolamento & purificação , Aeromonas/enzimologia , Aeromonas/genética , Aeromonas/isolamento & purificação , Proteínas de Bactérias/classificação , Proteínas de Bactérias/metabolismo , Brasil , Citrobacter/enzimologia , Citrobacter/genética , Citrobacter/isolamento & purificação , Enterobacter/enzimologia , Enterobacter/genética , Enterobacter/isolamento & purificação , Expressão Gênica , Humanos , Isoenzimas/genética , Isoenzimas/metabolismo , Klebsiella pneumoniae/enzimologia , Klebsiella pneumoniae/isolamento & purificação , Kluyvera/enzimologia , Kluyvera/genética , Kluyvera/isolamento & purificação , Pseudomonas/enzimologia , Pseudomonas/genética , Pseudomonas/isolamento & purificação , Recreação , Serratia/enzimologia , Serratia/genética , Serratia/isolamento & purificação , beta-Lactamases/classificação , beta-Lactamases/metabolismoRESUMO
Ornithine lipids (OLs) are phosphorus-free membrane lipids that can be formed by many bacteria but that are absent from archaea and eukaryotes. A function for OLs in stress conditions and in host-bacteria interactions has been shown in some bacteria. Some bacterial species have been described that can form OLs, but lack the known genes (olsBA) involved in its biosynthesis, which implied the existence of a second pathway. Here we describe the bifunctional protein OlsF from Serratia proteamaculans involved in OL formation. Expression of OlsF and its homologue from Flavobacterium johnsoniae in Escherichia coli causes OL formation. Deletion of OlsF in S. proteamaculans caused the absence of OL formation. Homologues of OlsF are widely distributed among γ-, δ- and ε-Proteobacteria and in the Cytophaga-Flavobacterium-Bacteroidetes group of bacteria, including several well-studied pathogens for which the presence of OLs has not been suspected, such as for example Vibrio cholerae and Klebsiella pneumonia. Using genomic data, we predict that about 50% of bacterial species can form OLs.
Assuntos
Aciltransferases/metabolismo , Lipídeos/genética , Lipídeos de Membrana/metabolismo , Ornitina/análogos & derivados , Serratia/enzimologia , Bacteroidetes/metabolismo , Cytophaga/metabolismo , Flavobacterium/metabolismo , Deleção de Genes , Lipídeos/biossíntese , Ornitina/biossíntese , Ornitina/genética , Proteobactérias/metabolismo , Serratia/metabolismoRESUMO
The increased incidence, high rates of mortality and few effective means of treatment of malignant melanoma, stimulate the search for new anti-tumor agents and therapeutic targets to control this deadly metastatic disease. In the present work the antitumor effect of arazyme, a natural bacterial-derived metalloprotease secreted by Serratia proteomaculans, was investigated. Arazyme significantly reduced the number of pulmonary metastatic nodules after intravenous inoculation of B16F10 melanoma cells in syngeneic mice. In vitro, the enzyme showed a dose-dependent cytostatic effect in human and murine tumor cells, and this effect was associated to the proteolytic activity of arazyme, reducing the CD44 expression at the cell surface, and also reducing in vitro adhesion and in vitro/in vivo invasion of these cells. Arazyme treatment or immunization induced the production of protease-specific IgG that cross-reacted with melanoma MMP-8. In vitro, this antibody was cytotoxic to tumor cells, an effect increased by complement. In vivo, arazyme-specific IgG inhibited melanoma lung metastasis. We suggest that the antitumor activity of arazyme in a preclinical model may be due to a direct cytostatic activity of the protease in combination with the elicited anti-protease antibody, which cross-reacts with MMP-8 produced by tumor cells. Our results show that the bacterial metalloprotease arazyme is a promising novel antitumor chemotherapeutic agent.