Behavior of deteriogenic fungi in aviation fuels (fossil and biofuel) during simulated storage.

Biofuels are expected to play a major role in reducing carbon emissions in the aviation sector globally. Farnesane ("2,6,10-trimethyldodecane") is a biofuel derived from the synthesized iso-paraffin route wich can be blended with jet fuel; however, the microbial behavior in farnesane/jet fuel blends remains unknown. The chemical and biological stability of blends should be investigated to ensure they meet the quality requirements for aviation fuels. This work aimed at evaluating the behavior of two fungi Hormoconis resinae (F089) and Exophiala phaeomuriformis (UFRGS Q4.2) in jet fuel, farnesane, and in 10% farnesane blend during simulated storage. Microcosms (150-mL flasks) were assembled with and without fungi containing Bushnell & Haas mineral medium for 28 days at a temperature of 20±2°C. The fungal growth (biomass), pH, surface tension, and changes in the fuel's hydrocarbon chains were evaluated. This study revealed thatthe treatment containing H. resinae showed a biomass of 19 mg, 12 mg, and 2 mg for jet fuel, blend, and farnesane respectively. The pH was reduced from 7.2 to 4.3 observed in jet fuel treatment The degradation results showed that compounds with carbon chains between C9 and C11, in jet fuel, and blend treatments were preferably degraded. The highest biomass (70.9 mg) produced by E. phaeomuriformis was in 10% farnesane blend, after 21 days. However, no significant decrease was observed on pH and surface tension measurements across the treatments as well as on the hydrocarbons when compared to the controls. This study revealed that farnesane neither inhibited nor promoted greater growth on both microorganisms.

Characterization of antimicrobial effect of organotin-based catalysts on diesel-biodiesel deteriogenic microorganisms.

Organotin compounds are applied in several industrial reactions and can present antifungal and antibacterial activities. Incorrect handling and storage practices of biodiesel and diesel-biodiesel blends can lead to microbial development, impacting its final quality. Concerning this problem, this work investigated the antimicrobial action of two organotin catalysts used in biodiesel production with four isolated microroorganisms (Bacillus pumilus, Pseudomonas aeruginosa, Pseudallescheria boydii, and Aureobasidium pullulans) and a pool of microorganisms (ASTM E1259 standard practice). Samples of soybean biodiesel with different concentrations of dibutyl tin dilaurate (catalyst 1) and di-n-butyl-oxo-stannane (catalyst 2) were prepared and added of mineral medium. The pool of microorganisms was inoculated and incubated at 30 °C and final biomass was weighted after 14 days. Thereafter, soybean biodiesel with catalyst 2 was used. Fungal biomass was weighted, and plate count was used to assess bacterial growth. Results show that catalysts 1 and 2 presented no inhibitory activity on the pool of microorganisms evaluated. A slight inhibitory activity was observed for B. pumilus and A. pullulans growth, but not for P. boydii, P. aeruginosa, or the pool of microorganisms. All experiment exhibited acidification higher than sterile control. Infrared analysis show less microbiological degradation products in the tin-protected fuel with ASTM inoculum. These results suggest that these tin-based catalysts show no toxic effect on native microbial population and a slight effect on some isolated microbial population in laboratory scale and for the first time shows that these organotin compounds can be employed safely as biodiesel catalyst. Graphical abstract.

Use of digital images to count colonies of biodiesel deteriogenic microorganisms.

This work presents a novel, robust procedure for the semi-automated counting of colony-forming units of Bacillus pumilus (a bacterium) and Meyerozyma guilliermondii (a yeast) both isolated from diesel oil. The counting is performed from digital images of Petri dishes containing the samples by a developed Python code, and the images are acquired from a low-cost scanning apparatus. The counting algorithm is based on the similar morphological characteristics of the bacterium and the yeast colonies. It was compared with classical counting methodology, and the results showed calibration and validation curves with a coefficient of determination (R2) of 0.99 and 0.98, respectively. The developed methodology is a valuable alternative to estimate the microbial contamination of biofuels.

Presence of antibiotic resistance genes and its association with antibiotic occurrence in Dilúvio River in southern Brazil.

It is known that antibiotics are widely used in human and veterinary medicine. In some countries the use is controlled, however few restrictions to their use are enforced in many countries. Antibiotics and their metabolites can reach the water bodies through sewage systems, especially in those countries with partial or absent wastewater treatment systems. The overuse and misuse of antibiotics has been linked with the increase of antibiotic resistant bacteria. The relation between the occurrence of antibiotics and resistance genes in surface waters has been widely studied worldwide evincing the great importance of this subject. In this work, a methodology for quantification of 40 antibiotics of 5 different classes, in river water, by SPE-LC-MS/MS was validated. Samples were taken during a two-year period from Dilúvio River, a stream that crosses the city of Porto Alegre (RS - Brazil) and receives in nature domestic effluent. The methodology met the requirements of validation, with Limit of Quantification varying from 20 ng L-1 to 100 ng L-1. A total of 48 samples was analyzed for the presence of antibiotics for two years. From the 40 antibiotics analyzed, 8 of them (Azithromycin, Cephalexin, ciprofloxacin, clindamycin, norfloxacin, sulfadiazine, sulfamethoxazole and trimethoprim) were present in all sampling points in the range of

Pseudallescheria boydii and Meyerozyma guilliermondii: behavior of deteriogenic fungi during simulated storage of diesel, biodiesel, and B10 blend in Brazil.

Due to their renewable and sustainable nature, biodiesel blends boost studies predicting their stability during storage. Besides chemical degradation, biodiesel is more susceptible to biodegradation due to its raw composition. The aim of this work was to evaluate the deteriogenic potential (growth and degradation) of Pseudallescheria boydii and Meyerozyma guilliermondii in degrading pure diesel (B0), pure biodiesel (B100), and a B10 blend in mineral medium during storage. The biodeterioration susceptibility at different fuel ratios and in BH minimal mineral medium were evaluated. The biomass measurements of P. boydii during 45 days indicated higher biomass production in the B10 blend. The growth curve of M. guilliermondii showed similar growth in B10 and B100. Although there was no significant production of biosurfactant, lipase production was detected in the tributyrin agar medium of both microorganisms. The main compounds identified in the aqueous phase by GC-MS were alcohols, esters, acids, sulfur, ketones, and phenols. The results showed that P. boydii grew at the expense of fuels, degrading biodiesel esters, and diesel hydrocarbons. M. guilliermondii grew in B100 and B10; however, degradation was not detected.

Draft Genome Sequence of Bacillus sp. Strain UFRGS-B20, a Hydrocarbon Degrader.

Bacillus sp. strain UFRGS-B20 was isolated in 2012 from Brazilian land-farming soil contaminated with petrochemical oily sludge. This strain was subjected to hydrocarbon biodegradation tests, showing degradation rates of up to 60%. Here, we present the 6.82-Mb draft genome sequence of the strain, which contains 2,178 proteins with functional assignments.

Metal resistance mechanisms in Gram-negative bacteria and their potential to remove Hg in the presence of other metals.

Contamination of the environment by heavy metals has been increasing in recent years due to industrial activities. Thus research involving microorganisms capable of surviving in multi-contaminated environments is extremely important. The objectives of the present study were to evaluate the removal of mercury alone and in the presence of cadmium, nickel and lead by four mercury-resistant microorganisms; estimate the removal of Cd, Ni and Pb; understand the mechanisms involved (reduction, siderophores, biofilms, biosorption and bioaccumulation) in the metal resistance of the isolate Pseudomonas sp. B50D; and determine the capacity of Pseudomonas sp. B50D in removing Hg, Cd, Ni and Pb from an industrial effluent. It was shown that the four isolates evaluated were capable of removing from 62% to 95% of mercury from a culture medium with no addition of other metals. The isolate Pseudomonas sp. B50D showed the best performance in the removal of mercury when evaluated concomitantly with other metals. This isolate was capable of removing 75% of Hg in the presence of Cd and 91% in the presence of Ni and Pb. With respect to the other metals it removed 60%, 15% and 85% of Cd, Ni and Pb, respectively. In tests with effluent, the isolate Pseudomonas sp. B50D removed 85% of Hg but did not remove the other metals. This isolate presented reduction, biosorption, biofilm production and siderophore production as its metal resistance mechanisms. Pseudomonas sp. B50D was thus a candidate with potential for application in the bioremediation of effluents with complex metal contaminations.

Mercury (II) removal by resistant bacterial isolates and mercuric (II) reductase activity in a new strain of Pseudomonas sp. B50A.

This study aimed to isolate mercury resistant bacteria, determine the minimum inhibitory concentration for Hg, estimate mercury removal by selected isolates, explore the mer genes, and detect and characterize the activity of the enzyme mercuric (II) reductase produced by a new strain of Pseudomonas sp. B50A. The Hg removal capacity of the isolates was determined by incubating the isolates in Luria Bertani broth and the remaining mercury quantified by atomic absorption spectrophotometry. A PCR reaction was carried out to detect the merA gene and the mercury (II) reductase activity was determined in a spectrophotometer at 340 nm. Eight Gram-negative bacterial isolates were resistant to high mercury concentrations and capable of removing mercury, and of these, five were positive for the gene merA. The isolate Pseudomonas sp. B50A removed 86% of the mercury present in the culture medium and was chosen for further analysis of its enzyme activity. Mercuric (II) reductase activity was detected in the crude extract of this strain. This enzyme showed optimal activity at pH 8 and at temperatures between 37 °C and 45 °C. The ions NH4(+), Ba(2+), Sn(2+), Ni(2+) and Cd(2+) neither inhibited nor stimulated the enzyme activity but it decreased in the presence of the ions Ca(2+), Cu(+) and K(+). The isolate and the enzyme detected were effective in reducing Hg(II) to Hg(0), showing the potential to develop bioremediation technologies and processes to clean-up the environment and waste contaminated with mercury.

Bioremediation assessment of diesel-biodiesel-contaminated soil using an alternative bioaugmentation strategy.

This study investigated the effectiveness of successive bioaugmentation, conventional bioaugmentation, and biostimulation of biodegradation of B10 in soil. In addition, the structure of the soil microbial community was assessed by polymerase chain reaction-denaturing gradient gel electrophoresis. The consortium was inoculated on the initial and the 11th day of incubation for successive bioaugmentation and only on the initial day for bioaugmentation and conventional bioaugmentation. The experiment was conducted for 32 days. The microbial consortium was identified based on sequencing of 16S rRNA gene and consisted as Pseudomonas aeruginosa, Achromobacter xylosoxidans, and Ochrobactrum intermedium. Nutrient introduction (biostimulation) promoted a positive effect on microbial populations. The results indicate that the edaphic community structure and dynamics were different according to the treatments employed. CO2 evolution demonstrated no significant difference in soil microbial activity between biostimulation and bioaugmentation treatments. The total petroleum hydrocarbon (TPH) analysis indicated a biodegradation level of 35.7 and 32.2 % for the biostimulation and successive bioaugmentation treatments, respectively. Successive bioaugmentation displayed positive effects on biodegradation, with a substantial reduction in TPH levels.

Isolation and characterization of bacteria from mercury contaminated sites in Rio Grande do Sul, Brazil, and assessment of methylmercury removal capability of a Pseudomonas putida V1 strain.

Methylmercury (MeHg) is one of the most dangerous heavy metal for living organisms that may be found in environment. Given the crescent industrialization of Brazil and considering that mercury is a residue of several industrial processes, there is an increasing need to encounter and develop remediation approaches of mercury contaminated sites. The aim of this study was to isolate and characterize methylmercury resistant bacteria from soils and sludge sewage from Rio Grande do Sul, Brazil. Sixteen bacteria were isolated from these contaminated sites and some isolates were highly resistant to methylmercury (>8.7 µM). All the isolates were identified by 16S rDNA. Pseudomonas putida V1 was able to volatilize approximately 90 % of methylmercury added to growth media and to resist to copper, lead, nickel, chromate, zinc, cobalt, manganese and barium. In the presence of high concentrations of methylmercury (12 µM), cell growth was limited, but P. putida V1 was still able to remove up to 29 % of this compound from culture medium. This bacterium removed an average of 77 % of methylmercury from culture medium with pH in the range 4.0-6.0. In addition, methylmercury was efficiently removed (>80 %) in temperature of 21-25 °C. Polymerase chain reactions indicated the presence of merA but not merB in P. putida V1. The growth and ability of P. putida V1 to remove methylmercury in a wide range of pH (4.0 and 8.0) and temperature (10-35 °C), its tolerance to other heavy metals and ability to grow in the presence of up to 11.5 µM of methylmercury, suggest this strain as a new potential resource for degrading methylmercury contaminated sites.

Capability of a selected bacterial consortium for degrading diesel/biodiesel blends (B20): enzyme and biosurfactant production.

The search for alternative sources of energy, such as biodiesel, has been stimulated, since this biofuel is highly susceptible for biodegradation and has low toxicity, thus, reducing the impact in ecosystems. The objective of this study was to select a bacterial consortium with potential for degrading diesel/biodiesel blends (B20) obtained from areas contaminated with hydrocarbons/esters. In order to evaluate the biodegrability of the blend, six enzyme assays were conducted: alkane hydroxylase, Catechol 1,2-dioxygenase, Catechol 2,3-dioxygenase, Protocatechol 3,4-dioxygenase, ρ-NPA hydrolysis (esterase), and release of fatty acids through titration (lipase), with estimative of total protein and biosurfactant production (surface tension measurement and emulsifying index E(24)). The best results obtained allowed the selection of four bacteria isolates (Bacillus megaterium, Bacillus pumilus, Pseudomonas aeruginosa, and Stenotrophomonas maltophilia) for compiling a consortium, which will be used for bioaugmentation strategies in soils contaminated with these fuels. This consortium exhibited high potential for biodegradation of biodiesel, and might be an efficient alternative for cleaning up these contaminated environments.

Propriedades químicas de um Argissolo tratado sucessivamente com composto de lixo urbano / Chemical properties of an Ultisol successively treated with municipal waste compost

Com o objetivo de avaliar as alterações em algumas propriedades químicas do solo sob adição sucessiva de composto de lixo urbano, conduziu-se um experimento a campo, em um solo Argissolo Vermelho de textura média na Estação Experimental Agronômica da UFRGS. Os tratamentos constaram de cinco doses de composto (0, 20, 40, 80 e 160t ha-1), de um tratamento com adubo mineral e uma testemunha (Calagem e NPK), com aplicações sucessivas por um período de quatro anos. A adição de composto no solo aumentou o pH, CTC, C orgânico, N total, P e Na extraíveis, bem como da relação de adsorção de sódio (RAS), Ca e Mg trocáveis e o teor de K extraível após a terceira aplicação. O Al trocável teve os seus teores diminuídos após aplicações sucessivas de composto. Os resultados permitiram concluir que aplicação de até 80t ha-1 por ano de composto de lixo urbano pode ser considerada como adequada para melhorar e, ou, manter as propriedades químicas do solo.

In order to assess the changes in some chemical properties of soil under the successive addition of urban waste compost, it was conducted a field experiment in an Ultisol soil at Agronomic Experimental Station of UFRGS. The treatments consisted of five successive applications of different doses of municipal solid waste compost (0, 20, 40, 80 and 160t ha-1) for a period of four years and a comparative treatment with mineral fertilizer and a control (without compost and fertilization). The addition of the compost increased soil pH, CEC, organic C, total N, extractable P and Na and the sodium adsorption ratio (SAR), calcium, and magnesium and extractable K after the third application. The exchangeable Al had their levels reduced by successive applications of compost. The results showed that application of up to 80t ha-1 yr-1 of urban waste compost can be the dose recommended to improve and maintain the chemical properties of soil.

Biorremediação de um solo contaminado com antraceno sob diferentes condições físicas e químicas / Bioremediation of a soil contaminated with anthracene under different chemical and physical conditions

O antraceno e os demais hidrocarbonetos aromáticos policíclicos (HAPs) podem ser removidos do solo pela biorremediação, cuja eficiência é limitada se as condições físicas e químicas não forem favoráveis à sobrevivência e à atividade dos microrganismos degradadores. O objetivo do presente estudo foi avaliar a influência do pH, da umidade e da disponibilidade de nitrogênio, de fósforo, de ferro e de enxofre na biorremediação de um solo contaminado com antraceno. Para tanto, amostras de um solo arenoso foram contaminadas em laboratório com 500mg kg-1 de antraceno e a mineralização desse poluente foi quantificada por respirometria. As maiores mineralizações ocorreram nos tratamentos com as maiores umidades e os pH avaliados. A adição de 100kg ha-1 ou mais de nitrogênio no solo e a redução da relação C HAP-N para valores inferiores a 120:17 diminuíram a mineralização do antraceno. O aumento da disponibilidade do fósforo, do ferro e do enxofre e a presença de amplas relações C HAP:P no solo não influenciaram a mineralização do antraceno. A correção do pH e o adequado fornecimento de água possibilitaram a biorremediação desse solo em curto período de tempo.

The anthracene, as well as the others polycyclic aromatic hydrocarbons (PAH), can be removed from the soil by bioremediation, whose efficiency is limited under unfavorable physical and chemical conditions to the survival and activity of the microbial degraders. The objective of this study was to evaluate the influence of pH, water content, and nitrogen, phosphorus, iron and sulfur concentrations in the bioremediation of a soil contaminated with anthracene. Samples of a sandy soil were contaminated in laboratory with anthracene (500mg kg-1) and the mineralization was evaluated by respirometry. The highest anthracene mineralization was verified in the soil with the highest water content and pH value studied. The addition of 100kg ha-1 nitrogen in the soil and the consequent reduction of CPAH-N ratio to values lesser than 120:17 reduced anthracene mineralization. The increase of phosphorus iron and sulfur availability and wide CPAH-P (120:1 to 120:22) ratios in the soil did not influence anthracene mineralization. The pH correction and appropriate water supply made possible the bioremediation of the soil polluted with anthracene in a short period of time.

Biorremediação de solos contaminados com hidrocarbonetos aromáticos policíclicos / Bioremediation of soils contaminated with polycyclic aromatic hydrocarbons

Os hidrocarbonetos aromáticos policíclicos (HAPs) são compostos mutagênicos e carcinogênicos aos humanos e aos animais que são introduzidos no ambiente em grandes quantidades, devido às atividades relacionadas à extração, ao transporte, ao refino, à transformação e à utilização do petróleo e de seus derivados. Apesar disso, a grande maioria dos microrganismos do solo não possui a capacidade de degradá-los, o que resulta na sua acumulação no ambiente e na conseqüente contaminação dos ecossistemas. Uma estratégia para a eliminação dos HAPs do solo é através da biorremediação, na qual os microrganismos que apresentam capacidade de metabolizar estes compostos irão transformá-los em substâncias inertes, CO2 e água. No entanto, esta biotecnologia pode ser limitada pela falta de microrganismos degradadores dos HAPs no solo, pela presença de condições ambientais desfavoráveis a estes microrganismos ou pela baixa biodisponibilidade dos contaminantes à microbiota degradadora. Para superar estas limitações e promover uma eficiente remoção dos contaminantes do ambiente, várias técnicas de biorremediação foram desenvolvidas, como biorremediação passiva, bioaumentação, bioestimulação, fitorremediação, landfarming, compostagem e biorreatores. Esta revisão visa a discutir os metabolismos bacteriano e fúngico destes compostos, os principais fatores químicos e físicos que influenciam a sobrevivência e a atividade destes microrganismos degradadores no ambiente e apresentar as técnicas de biorremediação que estão sendo atualmente utilizadas para a remoção dos HAPs no solo.

Polycyclic aromatic hydrocarbons (PAHs) are mutagenic and carcinogenic compounds to the humans and animals, released through the environment by anthropogenic activities related to the extraction, transport, refine, transformation and use of the petroleum and its derivatives. Most of the soils microorganisms do not possess the capacity to degrade them, which results in its accumulation in the atmosphere and contamination of the ecosystems. A strategy for PAHs elimination from the soil is through the bioremediation, where microorganisms having capacity to metabolize these compounds will transform them in inert substances, CO2 and water. However, this biotechnology can be limited by the lack of specific HAP microbial-degraders in soil, by unfavorable environmental conditions to these microorganisms or by the low bioavailability of those contaminants to the microorganisms. To overcome these limitations and to promote an efficient removal of the pollutants to the atmosphere, several bioremediation techniques were developed as passive bioremediation, bioaugmentation, biostimulation, phytoremediation, landfarming, composting and bioreactors. This revision aims at discussing microbial metabolism of PAHs, present the main chemical and physical factors that influence the survival and the activity of these microorganisms and to show the bioremediation techniques that are being used now for the PAHs removal in soil.

Bioremediation of soil contaminated by diesel oil

Avaliou-se a degradação de hidrocarbonetos de petróleo (HP) em solos contaminados com óleo diesel através da atenuação natural, bioestimulação e bioaumentação. A bioaumentação apresentou a maior degradação da fração leve (72,6%) e da fração pesada (75,2%) de HP e a atenuação natural foi mais efetiva do que a bioestimulação. A maior atividade da dehidrogenase no solo Long Beach e Hong Kong foi observada nos tratamentos bioaumentação e atenuação natural, respectivamente. O número de microrganismos degradadores de diesel e a população de heterotróficos não foi influenciada pelas técnicas de biorremediação. A melhor performance para a biorremediação do solo contaminado com diesel foi obtida quando foram adicionados microrganismos pré-selecionados do ambiente contaminado.

Bioremediation of soil contaminated by diesel oil

Were evaluated natural attenuation, biostimulation and bioaugmentation on the degradation of total petroleum hydrocarbons (TPH) in soils contaminated with diesel oil. Bioaugmentation showed the greatest degradation in the light (C12 - C23) fractions (72.7%) and heavy (C23 - C40) fractions of TPH (75.2%) and natural attenuation was more effective than biostimulation. The greatest dehydrogenase activity was observed upon bioaugmentation of the Long Beach soil (3.3-fold) and the natural attenuation of the Hong Kong soil sample (4.0-fold). The number of diesel oil degrading microorganisms and heterotrophic population was not influenced by the bioremediation treatments. The best approach for bioremediation of soil contaminated with diesel oil is the inoculum of microorganisms pre-selected from their own environment.

Avaliou-se a degradação de hidrocarbonetos de petróleo (HP) em solos contaminados com óleo diesel através da atenuação natural, bioestimulação e bioaumentação. A bioaumentação apresentou a maior degradação da fração leve (72,6%) e da fração pesada (75,2%) de HP e a atenuação natural foi mais efetiva do que a bioestimulação. A maior atividade da dehidrogenase no solo Long Beach e Hong Kong foi observada nos tratamentos bioaumentação e atenuação natural, respectivamente. O número de microrganismos degradadores de diesel e a população de heterotróficos não foi influenciada pelas técnicas de biorremediação. A melhor performance para a biorremediação do solo contaminado com diesel foi obtida quando foram adicionados microrganismos pré-selecionados do ambiente contaminado.