Bacterias degradadoras de Dimetilfenoles procedentes de aguas residuales

Motivación: Este estudio se centra en la biodegradación de compuestos fenólicos, como los dimetilfenoles, debido a los perjuicios causados en los seres humanos y en el medioambiente. Métodos: Se tomó muestras de aguas residuales procedentes de Leuna, cuyo municipio se encuentra la mayor industria química de Alemania, para aislar e identificar microorganismos bacterianos que degraden dimetilfenoles.La secuenciación de ADN y los análisis homólogos del gen ARNr 16S identificaron dos cepas; la cepa FGQ5 perteneciente a Delftia acidovorans y la cepa KNUC2106 perteneciente a Stenotrophomonas maltophilia.Ambas bacterias crecieron durante cuarenta y cinco días en cultivos líquidos con una mezcla de 3 isómeros de dimetilfenol (2,6-DMP, 3,4-DMP, 3,5-DMP), a una concentración de 50 mg/l, para adaptarlas a este medio como única fuente de carbono y posteriormente se inocularon, respectivamente, durante dos semanas en cada uno de los diferentes isómeros de dimetilfenol a una concentración de 70 mg/L. Resultados: Los valores experimentales indicaron que en la cepa FGQ5 se producía degradación de los isómeros 2,3-DMP y 3.4-DMP, mientras que no hubo valores positivos en el resto de isómeros en la fecha a la cuál fue tomada la última muestra. Por su parte la cepa KNUC2106 no mostró valores tangenciales que demostraran degradación de los diferentes dimetilfenoles, a fecha de la última muestra tomada.Conclusiones: Aunque la cepa KNUC2106 no mostrara valores positivos, presentaba un principio de cambio en cuanto a color y turbidez para los cultivos líquidos con isómeros 2,3-DMP y 3,4-DMP, respectivamente, que dejaba entrever la necesidad de más tiempo para que esta cepa bacteriana iniciara la degradación. Por otro lado la cepa FGQ5 degradó para los isómeros 2,3-DMP y 3.4-DMP pero se debe esperar más tiempo para comprobar si hay degradación en los restantes isómeros. En cuanto a futuros objetivos se debe realizar el mismo estudio a diferentes concentraciones de cada isómero de dimetilfenol para conocer la dosis máxima y mínima a las que pueden degradar ambas cepas bacterianas y por último amplificar el gen de la enzima catecol 2,3-dioxigenasa para comprobar su presencia como intermediadora en la degradación de dimetilfenoles

Degradation state of organic matter in surface sediments from the Southern Beaufort Sea: a lipid approach

For the next decades significant climatic changes should occur in the Arctic zone. The expected destabilisation of permafrost and its consequences for hydrology and plant cover should increase the input of terrigenous carbon to coastal seas. Consequently, the relative importance of the fluxes of terrestrial and marine organic carbon to the seafloor will likely change, strongly impacting the preservation of organic carbon in Arctic marine sediments. Here, we investigated the lipid content of surface sediments collected on the Mackenzie basin in the Beaufort Sea. Particular attention was given to biotic and abiotic degradation products of sterols and monounsaturated fatty acids. By using sitosterol and campesterol degradation products as tracers of the degradation of terrestrial higher plant inputs and brassicasterol degradation products as tracers of degradation of phytoplanktonic organisms, it could be observed that autoxidation, photooxidation and biodegradation processes act much more intensively on higher plant debris than on phytoplanktonic organisms. Examination of oxidation products of monounsaturated fatty acids showed that photo- and autoxidation processes act more intensively on bacteria than on phytodetritus. Enhanced damages induced by singlet oxygen (transferred from senescent phytoplanktonic cells) in bacteria were attributed to the lack of an adapted antioxidant system in these microorganisms. The strong oxidative stress observed in the sampled sediments resulted in the production of significant amounts of epoxy acids and unusually high proportions of monounsaturated fatty acids with a <i>trans</i> double bond. The formation of epoxy acids was attributed to peroxygenases (enzymes playing a protective role against the deleterious effects of fatty acid hydroperoxides in vivo), while <i>cis/trans</i> isomerisation was probably induced by thiyl radicals produced during the reaction of thiols with hydroperoxides. Our results confirm the important role played by abiotic oxidative processes in the degradation of marine bacteria and do not support the generally expected refractory character of terrigenous material deposited in deltaic systems

Degradación de compuestos aromáticos por la bacteria desnitrificante Magnetospirillum

Motivación: Se sabe que la bacteria Magnetospirillum sp. es degradadora de tolueno, pudiendo usar dicho compuesto como única fuente de carbono y energía. Es por esto que se quiere estudiar su capacidad para degradar otros compuestos que forman parte del petróleo y que están presentes en la vida diaria como materia prima para productos químicos como plásticos, pinturas, cosméticos, productos farmacéuticos y agroquímicos, los cuales son perjudiciales tanto para el hombre como para el medioambiente (1). Las sustancias que se ensayaron fueron los compuestos aromáticos fenol, p-cresol, benceno, tolueno, xileno así como benzoato de sodio, los cuales se han publicado como posibles candidatos para ser degradados por bacterias similares a la nuestra (2 y 3).Métodos: Una cepa de Magnetospirillum fue aislada desde un reactor de lecho fijo suministrado con  50 mg/L de tolueno. En el laboratorio, la cepa se mantuvo en un medio minimo (2), a pH 7.1 con tolueno, benzoato o succinato de sodio como única fuente de carbono. Los cultivos se incubaron a 30°C con agitación constante, en condiciones de anaerobiosis con KNO3 (5-10 mM) como aceptor de electrones. Los compuestos aromáticos que se ensayaron fueron fenol, p-cresol, benceno, xileno y tolueno a una concentración entre 0.05 mM a 1 mM. Se evaluó crecimiento bacteriano mediante espectrofotometría (560 nm) y se midió además concentración de tolueno mediante cromatografia de gases-FID. La presencia de genes catobólicos implicados en la ruta de degradación de tolueno, bssA y bcrC, fueron detectados mediante amplificación de genes por PCR utilizando cebadores especificos para Magnetospirillum sp.Resultados: La cepa fue capaz de degradar 50 mg/L de tolueno en 3 días y el máximo crecimiento bacteriano se alcanzó a esta concentración. Por el contrario, a 100 mg/L de tolueno, no se observó crecimiento significativo. A partir de densidad optica, se determinó que la cepa 15-1 crece además en succinato, benzoato y cresol. Los genes amplificados bssA y bcrC fueron detectados mediante la presencia de bandas de 188 pb y 159 respectivamente.Conclusiones: Si bien no se observó crecimiento bacteriano en varios de los compuestos ensayados, tolueno y p-cresol sí sustentaron el crecimiento bacteriano, demostrándose de esta manera que la capacidad para degradar está estrictamente limitado a la concentración y, por tanto, toxicidad de cada compuesto

Bacterias aerobias procedentes de humedales artificiales degradadoras de Dimetilfenoles

Motivación: Los humedales artificiales (HHAA) constituyen una tecnología apropiada para el tratamiento de las aguas residuales municipales e industriales. Estos sistemas tienen como papel principal la mineralización y transformación de los contaminantes orgánicos por los microorganismos que colonizan la rizosfera.  Los Dimetillfenoles (DMP) son componentes típicos de las aguas residuales de la industria petroquímica y carboquímica; tóxicos por ingestión y absorción de la piel [1].  Son compuestos peligrosos para la vida acuática, las plantas y muchos otros organismos, actuando como un sustrato inhibidor en la biotransformación [2]. Están presentes en 6 formas isómericas (DMP-2,3; DMP-2,4; DMP-2,5; DMP-2,6; DMP-3,4; DMP-3,5) [3]. Por esta razón, la comprensión de los procesos de degradación de bacterias que habitan en HHAA es un planteamiento importante para optimizar estos sistemas de tratamiento.Métodos: Diferentes HHAA fueron alimentados con aguas residuales fenólicas industriales y/o artificiales [4].  Las muestras de grava, agua y las raíces se inocularon en medio mínimo líquido con una mezcla equimolar de los isómeros DMP-2,6/3,4/3,5 como única fuente de carbono y de energía. Las bacterias fueron aisladas e identificadas por secuenciación del ADNr16S. Los aislados fueron inoculados en m.m con la anterior mezcla de isómeros de DMP, donde se obtuvo una única cepa degradadora. Ésta fue sometida a ensayos de degradación con cada uno de los isómeros por separado y con diferentes mezclas de isómeros.Resultados: De 20 aislados solo Delftia acidovorans fue capaz de degradar los isómeros de DMP-2,3 y DMP-3,4 por separados. La mezcla de DMP-2,3/2,5/3,4 resultó ser muy eficiente para nuestra bacteria, llegando todos los isómeros a una concentracion del 0%. También se observó que en la mezcla con los isómeros de DMP-2,6/3,4/3,5,  Delftia también degradó el isómero DMP-3,5.Conclusiones: D. acidovorans  es capaz de degradar solo los isómeros de DMP 2,3 y DMP-3,4, ya que resultan ser sustratos adecuados que proporcionan poder reductor y energía para la bacteria [5]. El isómero DMP-2,5, es solo degradado en presencia de los isómeros DMP-2,3 y DMP-3,4, al igual que el isómero DMP-3,5 que se degradada en la mezcla en presencia del isómero DMP-3,4. Probablemente se debe a un caso de cometabolismo, tratándose de una transformación colateral a la degradación del sustrato el cual induce la actividad enzimática para la degradación del cosustrato (DMP-2,5 y DMP-3,5) [6]

Two naphthalene degrading bacteria belonging to the genera Paenibacillus and Pseudomonas isolated from a highly polluted lagoon perform different sensitivities to the organic and heavy metal contaminants

Two bacterial strains were isolated in the presence of naphthalene as the sole carbon and energy source from sediments of the Orbetello Lagoon, Italy, which is highly contaminated with both organic compounds and metals. 16S rRNA gene sequence analysis of the two isolates assigned the strains to the genera Paenibacillus and Pseudomonas. The effect of different contaminants on the growth behaviors of the two strains was investigated. Pseudomonas sp. ORNaP2 showed a higher tolerance to benzene, toluene, and ethylbenzene than Paenibacillus sp. ORNaP1. In addition, the toxicity of heavy metals potentially present as co-pollutants in the investigated site was tested. Here, strain Paenibacillus sp. ORNaP1 showed a higher tolerance towards arsenic, cadmium, and lead, whereas it was far more sensitive towards mercury than strain Pseudomonas sp. ORNaP2. These differences between the Gram-negative Pseudomonas and the Gram-positive Paenibacillus strain can be explained by different general adaptive response systems present in the two bacteria

Two naphthalene degrading bacteria belonging to the genera Paenibacillus and Pseudomonas isolated from a highly polluted lagoon perform different sensitivities to the organic and heavy metal contaminants

Two bacterial strains were isolated in the presence of naphthalene as the sole carbon and energy source from sediments of the Orbetello Lagoon, Italy, which is highly contaminated with both organic compounds and metals. 16S rRNA gene sequence analysis of the two isolates assigned the strains to the genera Paenibacillus and Pseudomonas. The effect of different contaminants on the growth behaviors of the two strains was investigated. Pseudomonas sp. ORNaP2 showed a higher tolerance to benzene, toluene, and ethylbenzene than Paenibacillus sp. ORNaP1. In addition, the toxicity of heavy metals potentially present as co-pollutants in the investigated site was tested. Here, strain Paenibacillus sp. ORNaP1 showed a higher tolerance towards arsenic, cadmium, and lead, whereas it was far more sensitive towards mercury than strain Pseudomonas sp. ORNaP2. These differences between the Gram-negative Pseudomonas and the Gram-positive Paenibacillus strain can be explained by different general adaptive response systems present in the two bacteria

<i>Delftia</i> sp LCW, a strain isolated from a constructed wetland shows novel properties for dimethylphenol isomers degradation

BACKGROUND: Dimethylphenols (DMP) are toxic compounds with high environmental mobility in water and one of the main constituents of effluents from petro- and carbochemical industry. Over the last few decades, the use of constructed wetlands (CW) has been extended from domestic to industrial wastewater treatments, including petro-carbochemical effluents. In these systems, the main role during the transformation and mineralization of organic pollutants is played by microorganisms. Therefore, understanding the bacterial degradation processes of isolated strains from CWs is an important approach to further improvements of biodegradation processes in these treatment systems. RESULTS: In this study, bacterial isolation from a pilot scale constructed wetland fed with phenols led to the identification of Delftia sp. LCW as a DMP degrading strain. The strain was able to use the o-xylenols 3,4-DMP and 2,3-DMP as sole carbon and energy sources. In addition, 3,4-DMP provided as a co-substrate had an effect on the transformation of other four DMP isomers. Based on the detection of the genes, proteins, and the inferred phylogenetic relationships of the detected genes with other reported functional proteins, we found that the phenol hydroxylase of Delftia sp. LCW is induced by 3,4-DMP and it is responsible for the first oxidation of the aromatic ring of 3,4-, 2,3-, 2,4-, 2,5- and 3,5-DMP. The enzyme may also catalyze both monooxygenation reactions during the degradation of benzene. Proteome data led to the identification of catechol meta cleavage pathway enzymes during the growth on ortho DMP, and validated that cleavage of the aromatic rings of 2,5- and 3,5-DMPs does not result in mineralization. In addition, the tolerance of the strain to high concentrations of DMP, especially to 3,4-DMP was higher than that of other reported microorganisms from activated sludge treating phenols. CONCLUSIONS: LCW strain was able to degraded complex aromatics compounds. DMPs and benzene are reported for the first time to be degraded by a member of Delftia genus. In addition, LCW degraded DMPs with a first oxidation of the aromatic rings by a phenol hydroxylase, followed by a further meta cleavage pathway. The higher resistance to DMP toxicity, the ability to degrade and transform DMP isomers and the origin as a rhizosphere bacterium from wastewater systems, make LCW a suitable candidate to be used in bioremediation of complex DMP mixtures in CWs systems

Antimicrobial lubricant formulations containing poly(hydroxybenzene)-trimethoprim conjugates synthesized by tyrosinase

Poly(hydroxybenzene)-trimethoprim conjugates were prepared using methylparaben as substrate of the oxida- tive enzyme tyrosinase. MALDI-TOF MS analysis showed that the enzymatic oxidation of methylparaben alone leads to the poly(hydroxybenzene) formation. In the presence of tri- methoprim, the methylparaben tyrosinase oxidation leads poly(hydroxybenzene)-trimethoprim conjugates. All of these compounds were incorporated into lubricant hydroxyethyl cellulose/glycerol mixtures. Poly(hydroxybenzene)-trimetho- prim conjugates were the most effective phenolic structures against the bacterial growth reducing by 96 and 97 % of Escherichia coli and Staphylococcus epidermidis suspen- sions, respectively (after 24 h). A novel enzymatic strategy to produce antimicrobial poly(hydroxybenzene)-antibiotic conjugates is proposed here for a wide range of applications on the biomedical field.The authors Idalina Gonçalves and Cláudia Botelho would like to acknowledge the NOVO project (FP7-HEALTH- 2011.2.3.1- 5) for funding. Loïc Hilliou acknowledges the financial support by FCT – Foundation for Science and Technology, Portugal (501100001871), through Grant PEst-C/CTM/LA0025/2013 - Strategic Project - LA 25 - 2013–2014, and by Programa Operacional Regional do Norte (ON.2) through the project BMatepro – Optimizing Materials and Processes^, with reference NORTE-07-0124-FEDER-000037 FEDER COMPETE

Adaptation in toxic environments: Arsenic genomic islands in the bacterial genus Thiomonas:

Acid mine drainage (AMD) is a highly toxic environment for most living organisms due to the presence of many lethal elements including arsenic (As). Thiomonas (Tm.) bacteria are found ubiquitously in AMD and can withstand these extreme conditions, in part because they are able to oxidize arsenite. In order to further improve our knowledge concerning the adaptive capacities of these bacteria, we sequenced and assembled the genome of six isolates derived from the Carnoulès AMD, and compared them to the genomes of Tm. arsenitoxydans 3As (isolated from the same site) and Tm. intermedia K12 (isolated from a sewage pipe). A detailed analysis of the Tm. sp. CB2 genome revealed various rearrangements had occurred in comparison to what was observed in 3As and K12 and over 20 genomic islands (GEIs) were found in each of these three genomes. We performed a detailed comparison of the two arsenic-related islands found in CB2, carrying the genes required for arsenite oxidation and As resistance, with those found in K12, 3As, and five other Thiomonas strains also isolated from Carnoulès (CB1, CB3, CB6, ACO3 and ACO7). Our results suggest that these arsenic-related islands have evolved differentially in these closely related Thiomonas strains, leading to divergent capacities to survive in As rich environments